impacts of applied genetics: micro-organisms, plants, and animals

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Impacts of Applied Genetics: Micro-Organisms, Plants, and Animals

April 1981

NTIS order #PB81-206609



Library of Congress Catalog Card Number 81-600046

For sale by the Sup erintendent of Documents,U.S. Governmen t Printing Office, Wash ington, D.C. 20402



F o r e w o r d

This report examines the ap plication of classical and molecular genetic technol-ogies to micro-organisms, plants, and animals. Congressional sup port for an assess-ment in the field of genetics dates back to 1976 when 30 Representatives requ ested astudy of recombinant DN A technology. Letters of sup port for this broader stu dy camefrom the then Senate Committee on Human Resources and the House Committee onInterstate and Foreign Commerce, Subcomm ittee on H ealth and the Environment.

Current developm ents are especially rapid in the application of genetic technol-ogies to micro-organisms; these were studied in three industries: pharmaceutical,chemical, and food. Classical genetics continue to play the major role in plant andanimal breeding but n ew genetic techniques are of ever-increasing importance.

This report id entifies and d iscusses a number of issues and options for the Con-gress, such as:

q Federal Government support of R&D,q methods of improving the germplasm of farm animal species,. risks of genetic engineering,

patenting living organisms, andq pu blic involvement in decisionmaking.

The Office of Technology Assessment was assisted by an advisory panel of scien-tists, industrialists, labor representatives, and scholars in the fields of law, economics,and those concerned with the relationships betw een science and society. Others con-tributed in two w orkshop s held du ring the course of the assessment. The first was toinvestigate pu blic perception of the issues in genetics; the second examined geneticapplications to animals. Sixty reviewers drawn from universities, Government, in-du stry, and the law provided helpful comm ents on d raft reports. The Office expressessincere ap preciation to all those ind ividu als.

An abbreviated copy of the summary of this report (ch. 1) is available free of charge from the Office of Technology Assessment, U.S. Congress, Washington, D. C.,20510. In ad dition, the working p apers on the u se of genetic technology in hu man andin veterinary med icine are available as a sep arate volume from the N ational TechnicalInformation Service.

Director



Im p ac t s o f App l ied Gen e t i c s Adv i so ry

J. E. Legates , ChairmanDean, School of Agriculture and Life Sciences, North Carolina State University

Ronald E. CapeCetus Corp.

Nina V. Fedoroff

Department of EmbryologyCarnegie Institution of Washington

June GoodfieldThe Rockefeller University

Harold P. GreenFried, Frank, Harr is, Shriver an d Kamp elman

Halsted R. HolmanStanford University Med ical School

M. Sylvia KrekelHealth and Safety OfficeOil, Chemical, and Atomic Workers

International UnionElizabeth Kutter

The Evergreen State College

Oliver E. Nelson, Jr.Laboratory of GeneticsUniver sity of Wisconsin

David PimentelDepartm ent of EntomologyCornell University

Robert WeaverDepartm ent of Agricultural EconomicsPennsylvania State Un iversity

James A. WrightPioneer H i-Bred InternationalPlant Breeding Division

Norton D. ZinderThe Rockefeller University

iv



App l ied Gen e t i c s Asses smen t S ta ff

Joyce C. Lashof, Assistant Director, OTA Health and Life Sciences Division

Gretchen Kolsrud , Program Manager

Zsolt Harsanyi, Project Director

Project Staff

Marya Breznay, Administrative AssistantLawrence Burton , Analyst

Susan Clymer, Research AssistantRenee G. Ford, * Technical EditorMichael Gough, Senior AnalystRobert Grossman,* Analyst

Richard H utton, * EditorGeoffrey M. Karny, Legal Analyst

Major Contractors

Benjamin G. Brackett, University of Penn sylvan iaThe Genex Corp.William P. O’Neill, Poly-Planning ServicesPlant Resources InstituteAnthony J. Sinskey, Massachusetts Institute of Technology

Aladar A. Szalay, Boyce-Thompson InstituteVirginia Walbot, Washington University

OTA P u b l i sh in g S t a f f

John C. Holmes, publishing Officer

John Bergling* Kathie S. Boss Debra M. Datcher

Patricia A. Dyson* Mary Harvey * Joe Hen son

OTA contract personnel.

v



C o n t e n t s

Page

Glossary . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. .. .viii. . .

Acronyms and Abbreviations . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . xii

Chapter l.

Chapter 2.

Chapter 3.

Chapter 4.

Chapter 5.

Chapter 6.

Chapter 7.

Summary: Issues and Options . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3

Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .......00 29

Part I :Biotechnology

Genetic Engineering and the Fermentation Technologies . . . . . . . . . . 49

The Pharmaceutical Industry. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 59

The Chemical Industry . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 85

The Food Processing Industry. . . . . . . . . . . . . . . . . . . . . . . . . .. ....107

The Use of Genetically Engineered Micro-Organisms in theEnvironment. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . ......117

Part II : Agriculture

Chapter 8. The Application of Genetics to Plants . . . . . . . . . . . . . . . . . . . . .. ...137Chap ter 9. Ad vances in Reprod uctive Biology and Their Effects on

Animal Improvement . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 167

Chapter l0.

Chapter ll.

Chapter 12.

Chapter 13.

Appendixes

Part III: Institutions and Society

The Question of Risk. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .......199

Regulation of Genetic Engineering. . . . . . . . . . . . . . . . . . . . . .. ....211

Patenting Living Organisms. . . . . . . . . . . . . . . . . . . . . . ..........237

Genetics and Society . . . . . . . . . . . . . . . . . . . . . ................257

I-A. A Case Study of Acetaminophen Production. . . . . . . . . . . . . . . . . . . . . ...269I-B. A Timetable for the Commercial Production of Compounds Using

Genetically Engineered Micro-Organisms in Biotechnology . . . . ........275

I-C. Chemical and Biological Processes. . . . . . . . . . . . . . . . . . . . . . ..........292

I-D. The Impact of Genetics on Ethanol-A Case Study . . . . . . .............293

II-A. A Case Study of Wheat. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. ...304

11-B. Genetics and the Forest Products Industry Case Study . . . . . . . .........307

II-C. Animal Fertilization Technologies. . . . . . . . . . . . . . . . . . . . . . ..........309

III-A. History of the Recombinant DNA Debate . . . . . . . . . . . . . . . . . . . . .. ...315

III-B. Constitutional Constraints on Regulation. . . . . . . . . . . . . . . . . . . . . .. ...320

111-C. Information on International Guidelines for Recombinant DNA. . ......322IV. Planning Workshop Participants, Other Contractors and Contributors, and

Acknowledgments. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . ...329

vii



G l o ssa r y

Aerobic.—Growing on ly in th e presence of oxygen.

Anaerobic.—Growing only in the absence of oxygen.

Alkaloids.—A group of nitrogen-containing organic

substances found in p lants; many are ph armaco-logically active—e.g., nicotine, caffeine, andcocaine.

Allele.—Alternate form s of th e same gene. For ex-ample, the genes responsible for eye color (blue,brown, green, etc.) are alleles.

Amino acids.—The building blocks of proteins.There are 20 common amino acids; they are’ joined together in a strictly ordered “string”which determines the character of each protein.

Antibody.—A protein component of the immun esystem in mammals found in the blood.

Antigen.—A large molecule, usually a protein orcarbohydrate, which when introduced in thebody stimulates the production of an antibodythat will react specifically with the antigen.

Aromatic chemical.—An organic compound con-taining one or more six-membered rings.

Aromatic polymer.–Large molecules consistingof repeated structural units of aromatic chem-icals.

Artificial insemination.—The manual placementof sperm into the uterus or oviduct.

Bacteriophage (or phage).–A virus that multi-plies in bacteria. Bacteriophage lambda is com-monly used as a vector in recombinant DNA ex-periments.

Bioassay .–Determin ation of th e relative strengthof a substance (such as a drug) by comparing itseffect on a test organism with that of a standardpreparation.

Biomass.—Plant and animal material.

Biome.—A community of living organisms in a major ecological region .

Biosynthes is . —The production of a chemical com-

poun d by a living organism.

Biotechnology .—The collection of industrial proc-esses that involve the use of biological systems.For some of these industries, these processes in-

volve the use of genetically engin eered micro-organisms.

Blastocyst.— An early developmental stage of theembryo; the fertilized egg undergoes several celldivisions and forms a hollow ball of cells calledthe blastocyst.

Callus.—The cluster of plant cells that results fromtissue culturing a sin gle plant cell.

Carbohydrates.— The family of organic moleculesconsisting of simple sugars such as glucose andsucrose, and su gar chains (polysaccharides) su chas starch and cellulose.

Catalyst.—A substance that enables a chemicalreaction to take place under milder than normalconditions (e.g., lower temperatures). Biologicalcatalysts are enzymes; nonbiological catalysts in-clude metallic complexes.

Cell fusion.—The fu sing together of two or m orecells to become a single cell.

Cell lysis.—Disruption of the cell membrane allow-ing the breakdown of the cell and exposure of itscontents to the environment.

Cellulase.—An en zyme that d egrades cellulose toglucose.

Cellulose.–A polysaccharid e composed entirely of several glucose units linked end to end; it consti-tutes the major part of cell walls in plants.

Chimera.—An individual composed of a mixture of

genetically different cells.

Chloroplast.-The structure in plant cells wherephotosynthesis occurs.

Chromosomes.—The thread-like components of acell that are composed of DNA and protein. Theycontain most of th e cell’s DN A.

Clone.—A group of genetically identical cells ororganisms asexually descended from a commonancestor. All cells in the clone have th e same ge-netic material and are exact copies of the original.

Conjugation.—The one-way transfer of DN A be-tween bacteria in cellular contact.

Crossing-over. —A genetic event that can occurduring celluar replication, which involves thebreakage and reun ion of DNA molecules.

Cultivar.—An organism developed and persistentunder cultivat ion.



Cytognetics.-A branch of biology that deals withthe study of heredity and variation by the meth-ods of both cytology (the study of cells) andgenetics.

Cytoplasm.—The p rotoplasm of a cell, external tothe cell’s nuclear membrane.

Diploid.—A cell with double the basic chromosomenumber.

DNA (deoxyribonucleic acid).—The genetic ma-terial found in all living organisms. Every inher-ited characteristic has its origin somewh ere inthe code of each ind ividual’s complement of D NA.

DNA vector.—A vehicle for transferring DNA fromone cell to another.

Dominant gene.—A characteristic whose expres-sion prevails over alternative characteristics for agiven trait.

Escherichia coli.-A bacterium that commonly in-habits the human intestine. It is a favorite orga-nism for many microbiological experiments.

Endotoxins.-Complex molecules (lipop olysaccha-rides) that compose an in tegral part of the cellwall, and are released only when the integrity of the cell is disturbed.

Embryo t ransfer .— Implantation of an embryointo the oviduct or uterus.

Enzyme.—A functional protein that catalyzes achemical reaction. Enzymes control th e rate of metabolic processes in an organism; they are theactive agents in the fermentation process.

Estrogens.—Female sex hormones.

Estrus (“heat").-The period in which the female

will allow the male to mate her.

Euk aryote.—A higher, compartmentalized cellcharacterized by its extensive internal structureand the presence of a nucleus containing theDNA. All multicellular organisms are eukaryotic.The simp ler cells, the p rokaryotes, have muchless compartmentalization and internal struc-ture; bacteria are prokaryotes.

Exotoxins.—Proteins produced by bacteria that areable to diffuse out of the cells; generally more po-tent and specific in their action than endotoxins.

Fermentation.—The biochemical process of con-v

er

tin

g a raw material such as glucose into a

product such as ethanol.

Fibroblast.—A cell that gives rise to connectivetissues.

Gamete.—A mature reproductive cell.

Gene.–The hereditary unit; a segment of DNAcoding for a specific protein.

Gene expression.—The manifestation of the ge-netic material of an organism as specific traits.

Genetic drift.—Changes of gene frequency in smallpopulation due to chance preservation or extinc-tion of p articular genes.

Gen etic code.—The b iochem ical basis of heredityconsisting of codons (base triplets along th e DN Asequence) that determine the specific amino acidsequence in proteins and that are the same for allforms of life studied so far.

Genetic engineering.-A technology used at thelaboratory level to alter the hereditary apparatusof a living cell so that the cell can produce moreor different chemicals, or perform completelynew functions. These altered cells are then usedin industrial production.

Gene mapping-Determining the relative loca-tions of different genes on a given chromosome.

Genome.—The bas ic chromosome set of an

organism—the su m total of its genes.

Genotype.—The genetic constitution of an individ-ual or group.

Germplasm.-The total genetic variability availableto an organism, represented by the pool of germcells or seed.

Germ cell.—The sex cell of an organ ism (sperm oregg, pollen or ovum). It differs from other cells inthat it contains only half the usu al num ber of chromosomes. Germ cells fuse during fertiliza-tion.

Glycopeptides.—Chains of amino acids with at-tached carbohydrates.

Glycoprotein.— A conjugated protein in which thenonprotein group is a carbohydrate.

Haploid .—A cell with only one set (half of the usualnumber) of chromosomes.

Heterozygous.—When the two genes controlling aparticular trait are different, the organism isheterozygous for that trait.

Homozygous.— When the two genes controlling aparticular trait are iden tical for a p air of chro-mosomes, the organism is said to be homozygous

for that trait.

Hormones.—The “messenger” molecules of thebody that help coordinate the actions of varioustissues; they produce a specific effect on the ac-tivity of cells remote from their point of origin.

ix



Hybrid.—A new variety of plant or animal that re- Nif genes.—The genes for nitrogen fixation presentsuits from cross-breeding two different existing in certain bacteria.varieties.

Nucleic acid.—A polymer composed of DNA orHyd rocarbon . —All organic compounds that are RNA subunits .

composed on ly of carbon an d h ydrogen.Nucleotides.—The fundamental units of nucleic

Im m unopro te ins .— All the proteins that are part acids. They consist of one of the four bases—of the immu ne system (including antibodies, in- adenine, guanine, cytosine, and thymine (uracilterferon, and cytokines). in the case of RNA)—and its attached sugar-phos-

In vitro. -outside the living organism and in anphate group.

artificial environment. O r g an i c co mp ou n d s.—Ch em ical compounds

In vivo.—Within the living organism.based on carbon chains or rings, which containhyd rogen, and also may contain oxygen, nitro-

Leukocytes.—The white cells of blood. gen, and various other elements.

Lipids.–Water insolub le biomolecules, such as cel- Parthenogenesis.— Reproduction in animals with-lular fats and oils. out male fertilization of the egg.

Lipopolysaccharides. —Complex substances com- Pathogen.—A specific causative agent of d isease.posed of lipids and p olysaccharides.

Peptide. —Short chain of amino acids.L y m p h o b l a s t o i d . – Referring to malignant white

blood cells.pH .–A measure of the acidity or b asicity of a solu-

tion; on a scale of O (acidic) to 14 (basic): for exam-Lymphokines.–The biologically active soluble fac-

tor produced by w hite blood cells.

Maleic anhydride.—An important organic chem-ical used in the manufacture of synthetic resins,in fungicides, in the dyeing of cotton textiles, andto prevent the oxidation of fats and oils duringstorage and rancidity.

Messenger RNA.–Ribonucleic acid molecules thatserve as a guide for protein synthesis.

Metabolism.—The sum of the physical and chem-ical processes involved in the m aintenance of lifeand b y which energy is made available.

Mitochondria.—Structures in higher cells thatserve as the “powerhouse” for the cell, producingchemical energy.

Monoclinal antibodies.—Antibodies derivedfrom a single source or clone of cells whichrecognize only one kin d of antigen.

Mutants.–Organisms whose visible properties withrespect to some trait differ from the norm of thepopulation due to mutat ions in i ts DNA.

pie, lemon juice has a pH of 2.2 (acidic), water hasa pH of 7.0 (neu tral), and a solution of bak ing

soda h as a p H of 8.5 (basic).

Phage.—(See bacteriophage.)

Phenotype. —The visible properties of an organismthat are produced by the interaction of the geno-type and the environment.

Plasmid.—Hereditary material that is not p art of achromosome. Plasmids are circular and self-repli-cating. Because they are generally small and rela-tively simple, they are used in recombinant DNAexperiments as acceptors of foreign DNA.

Plastid.–Any specialized organ of the plan t cell

other than the nucleus, such as the chloroplast.Ploidy. —Describes the nu mber of sets of chromo-

somes present in the organism. For example,humans are d iploid, having two homologous setsof 23 chromosom es (one set from each parent)for a total of 46 chromosomes; many p lants arehaploid, having only one copy of each chro-mosome.

Mutation.—Any change that alters the sequence of Polymer.—A long-chain molecule formed from

bases along the DNA, changing the genetic ma- smaller repeating structural units.

terial. Polysacchar ide . —A long-chain carbohydrate con-

Myeloma.—A malignant disease in which tumor taining at least three molecules of simple sugars

cells of the antibody producing system synthesize linked together; examples would include cellu-excessive amounts of specific proteins. lose and starch.

n-alkanes.—Straight chain h yd rocarb on s–th e Pr og est og en s. —Hormones involved w ith ovula-main constituents of petroleum. tion.

x



Prostaglandin.-Refers to a group of naturally oc-curring, chemically related long-chain fatty acidsthat have certain physiological effects (stimulatecontraction of uterine and other smooth muscles,lower blood pressure, affect action of certainhormones).

Protein.—A linear polymer of amino acids; proteinsare the products of gene expression and are thefunctional and structural components of cells.

Protoplasm.—A cell without a wall.

Protoplasm fusion.—A means of achieving genetictransformation b y joining two protoplasts or join-ing a protoplasm with any of the components of another cell.

Recess ive gene. —Any gene whose expression isdependent on the absence of a dominant gene.

Recombinant DNA.–The hybrid DNA prod ucedby joining pieces of DNA from different sources.

Restriction enzyme. —An enzym e within a bac-terium that recognizes and degrades DNA fromforeign organisms, thereby preserving the genet-

ic integrity of the bacterium. In recombinantDNA experiments, restriction enzymes are usedas tiny biological scissors to cut u p foreign D NAbefore it is recombined with a vector.

Reverse transcriptase.—An enzyme that can syn-thesize a single strand of DNA from a messenger

RNA, the reverse of the normal direction of proc-essing genetic information within the cell.

RNA (ribonucleic acid).—In its three forms—mes-senger RNA, transfer RNA, and ribosomal RNA—it assists in translating the genetic message of DNA into the finished protein .

Somatic cell.—One of th e cells comp osing p arts of the body (e.g., tissues,cell.

Tissue culture.—An ining healthy cells fromfrom skin.

organs) other than-a germ

vitro method of p ropagat-tissues, such as fibroblasts

Transduction.—The process by which foreignDNA becomes incorporated into the genetic com-plement of the host cell.

Transformation.—The transfer of genetic infor-mation by DNA separated from the cell.

Vector.—A transmission agent; a DNA vector is aself-replicating DN A molecule that transfers apiece of DNA from one h ost to another.

Virus.–An infectious agent that requires a host cellin order for it to replicate. It is composed of either RNA or DNA wrapped in a protein coat.

Zygote.—A cell formed by the union of two maturereproductive cells.

xi



Ac r o n y m s a n d Ab b r e v ia t i o n s

AAAC SACTHAIAIPL

APAPAS Mbblbbl/ dBOD5BBMbuCaMVCCPA

CD CCERB

D H H S

D H I

D N AD O DD O DDOED P A GEOREPAFD AFMDVft z

ft

FTC

gga lG Hh aH EW

h G HH YV

— amin o acids— American Cancer Society— adrenocorticotropic hormone— artificial insemination— Animal Improvement Programs

Laboratory

— acetaminoph en— American Society for Microbiology– barrel(s)– barrels per day— 5-day biochemical oxygen demand— Biological Response Modifier Program– bushel— cauliflower mosaic virus— The Court of Customs and Patent

Appeals— Center for Disease Control— Cambridge Experimentation Review

Board— Department of Health and Human

Services (formerly Health, Education,and Welfare)

— Dairy Herd Improvement– deoxyribonucleic acid— Department of Commerce— Department of Defense— Department of Energy— Dangerous Pathogens Advisory Group— enhanced oil recovery— Environmental Protection Agency— Food and Drug Administration— foot-and-mouth disease virus— square foot– foot— Federal Trade Commission– gram– gallon– growth h ormone— hectares— Department of Health, Education, and

Welfare– hum an growth hormone— high-yielding varieties

IBCs — Institutional Biosafety CommitteesIC I — Imperial Chemical IndustriesIN D — Investigational New Drug Application

(FDA)k g – kilogram1 — liter

lb – poun dm g — m illigram

1% — microgrampm — micrometer (formerly micron)MUA — Memorandum of Understanding and

AgreementNCI — National Cancer InstituteN D A — new drug application (FDA)N D A B — National Diabetics Advisory BoardN D C H IP — National Cooperative Dairy Herd

ProgramNIAID — National Institute of Allergy and

Infectious DiseasesNIAMDD — National Institute of Arthritis,

Metabolism, and Digestive DiseasesN IH – National Institutes of HealthNIOSH — National Institute of Occupational

Safety and HealthN SF — National Science FoundationOECD — The Organization for Economic

Cooperation and DevelopmentO R D A — Office of Recombinant DNA ActivitiesPD — predicted differencep H — unit of measure for acidity/ basicityp p m – parts per millionR& D — research and developmentRA C — Recombinan t DN A Advisory CommitteerDNA — recombin ant DN A

SC P — single-cell proteinT-DNA — a smaller segment of the the p lasmidTi — tumor inducingTSCA — Toxic Substances Control Act



ch a p t e r 1

Su m m a r y: I ssu e s a n d O p t i on s



C h a p t e r 1

Page

Biotechnology . . . . . . . . . . . . . . . . . . . . . . . . . . . 4

The Pharmaceutical industry. . . . . . . . . . . . . . 4Findings ... ... ..... . . . . . . . . . . . . . . . . 4

The Chemical Industry. . . . . . . . . . . . . . . . . . . 7Findings . . . . . . . . . . . . . . . . . . . . . . . . . . . . 7

Food processing industry. . . . . . . . . . . . . . . . . 8Findings . . . . . . . . . . . . . . . . . . . . . . . . . . . . 8

The Use of Genetically Engineered Micro-Organ isms in the Environ men t . . . . . . . . . . . 8Findings . . . . . . . . . . . . . . . . . . . . . . . . . . . . 8Min eral Leachin g an d Recovery . . . . . . . . . . 9Enhanced Oil Recovery. . . . . . . . . . . . . . . . . 9Pollution Control . . . . . . . . . . . . . . . . . . . . . 9Constraints in Using Genetic EngineeringTechn ologies in O pen Environ men ts. . . . . . . 10

Issue and Options—Biotechnology . . . . . . . . . . 10Agriculture . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 11

The Ap plications of G enetics to Plants . . . . . . . 11

Findings . . . . . . . . . . . . . . . . . . . . . . . . . . . . 11

New Genetic Technologies for Plant Breeding 12Constraints on Using Molecular Gen etics

for Plant Improvements. . . . . . . . . . . . . . . 13Genetic Variability, CropVulnerability, and

the Storage of Germplasm . . . . . . . . . . . . . 13Issu es an d Op tion s—Plan ts. . . . . . . . . . . . . . . . 14Advances in Reprodu ctive Biology and Their

Effects on Anim al Im prov emen t . . . . . . . . . . 15

Page

Findings . . . . . . . . . . . . . . . . . . . . . . . . . . . . 17Issue and Options-Animals . . . . . . . . . . . . . . . 17

Institu tion s an d So ciety. . . . . . . . . . . . . . . . . 18

Regulation of Genetic Engineering . . . . . . . . . . 18Findings . . . . . . . . . . . . . . . . . . . . . . . . . . . . 18

Issue and Op tion s—Regulation . . . . . . . . . . . . . 20Paten tin g Livin g O rgan ism s . . . . . . . . . . . . . . . 22

Findings . . . . . . . . . . . . . . . . . . . . . . . . . . . . 22Issue and Options —Patenting Living Organisms 23Genetics and Society . . . . . . . . . . . . . . . . . . . . 24Issues and options—Genetics and Society. .. . 24

TableTable No. Page

1. Containment Recommended by the NationalInstitutes of Health... . . . . . . . . . . . . . . . . . . 19

FiguresFigure No. Page

1. Recombinant DNA: The Technique of Recombinin g Genes From On e Species WithThose From Another.. . . . . . . . . . . . . . . . . . . 5

2. The Pro du ct Develop men t Process . . . . . . . . . 63. The Way the Reproductive Technologies

Interrelate . . . . . . . . . . . . . . . . . . . . . . . . . . . . 16



Ch ap te r 1

S u m m a r y : I ss u e s a n d O p t io n s

The genetic alteration of plants, animals, and

micro-organisms has been an important part of

agriculture for centuries. It has also been an in-tegral part of the alcoholic beverage industry

since the invention of beer and wine; and for

the past century, a mainstay of segments of the

pharmaceutical and chemical industries.

However, only in the last 20 years have pow-

erful new genetic technologies been developed

that greatly increase the ability to manipulate

the inherited characteristics of plants, animals,

and micro-organisms. One consequence is the

inc reas ing r e l i ance the pha rm aceu t i ca l and

chemical industries are placing on biotechnol-

ogy. Micro-organisms are being used to manu-facture substances that have previously beenextracted from natural sources. Animal andplant breeders are using the new techniqu es tohelp clarify basic questions about biologicalfunctions, and to improve the speed and effi-ciency of the technologies they already use.Other ind ustries–from food p rocessing an d pol-lution control to m ining and oil recovery—areconsidering the use of genetic engineering to in-crease p rodu ctivity and cut costs.

Genetic technologies will have a broad impact

on the future. They may contribute to fillingsome of the most fund amental needs of man-kind–from health care to sup plies of food andenergy. At the same time, they arouse concernsabout their potential effects on the environmentand the risks to health involved in basic andapplied scientific research and development(R&D). Because genetic technologies are alread ybeing applied, it is appropriate to begin con-sidering their potential consequences.

Congressional concern with applied geneticsdates back to 1976, when 30 Representatives re-quested an assessment of recombinant DNA(rDNA) technology. Support for the broaderstudy reported here came in letters to the Officeof Technology Assessment from the then SenateCommittee on Human Resources and the HouseCommittee on Interstate and Foreign Com-merce, Subcomm ittee on Health and the Envi-

spronm ent. In add ition specific subtop ics are of interest to other committees, notably those hav-

ing jurisdiction over science and technology andthose concerned w ith patents.

This report describes the potentials and prob-lems of applying the new gen etic technologies toa range of major industries. It emphasizes thepresent state of the art because that is whatdefines the basis for the futu re applications. Itthen makes som e estimates of econom ic, envi-ronmental, and institutional impacts—where,when, and how some technologies might be ap-plied and what some of the results might be.The report closes with the possible roles that

Government, industry, and the public mightplay in determining the future of appliedgenetics.

The term applied genetics, as used in thisreport, refers to two groups of technologies:

q Classical genetics—natural mating methodsfor the selective breeding of organismsfor desired characteristics–e. g., breedingcows for increased milk production. Thepool of genes available for selection is com-prised of those that cause natural differ-ences among individuals in a population

and those obtained by m utation.q Molecular genetics includes the technologies

of genetic engineering that involve thedirected manipulation of the genetic mate-rial itself. These technologies—such asrDNA and the chemical synthesis of genes—can increase the size of the gene pool forany one organism by making available ge-netic traits from many different popula-tions. Molecular genetics also includestechnologies in which manipulation occursat a level higher th an that of the gene—atthe cellular level, e.g., cell fusion and invitro fertilization.

Significant applications of molecular geneticsto micro-organisms, such as the efforts to man-ufacture hu man insulin, are already un derwayin several industries. Most of these applications

3



4 q Impacts of Applied Genetics—Micro-Organisms, Plants, and Animals

dep end on fermentation—a technology in w hich plants an d animals, which are biologically m oresubstances produced by micro-organisms can complex and more difficult to manipulate suc-be obtained in large quan tities. Applications to cessfully, will take longer to develop.

Bio techno logy

Biotechnology-the use of living organisms ortheir components in industrial processes—ispossible because micro-organisms naturally pro-duce countless substances during their lives.Some of these substances have proved commer-cially valuable. A number of different industrieshave learned to use micro-organisms as naturalfactories, cultivating populations of the bestproducers under conditions designed to en-hance their abilities.

App lied genetics can p lay a m ajor role in im-

proving the sp eed, efficiency, and p rodu ctivityof these biological systems. It permits the ma-nipulation, or engineering, of the micro-orga-nisms’ genetic material to produce the d esiredcharacteristics. Genetic engineering is not initself an industry, but a technique used at thelaboratory level that allows the researcher tomod ify the h ereditary app aratus of the cell. Thepop ulation of altered identical cells that grow sfrom the first changed micro-organism is, inturn, used for various industrial processes. (Seefigure 1.)

The first major commercial effects of the ap-plication of genetic engineering will be in theph armaceutical, chemical, and food p rocessingindustries. Potential commercial applications of value to the m ining, oil recovery, and pollutioncontrol industries—wh ich m ay d esire to use m a-nipulated m icro-organisms in the open environ -ment —are stll somewhat speculative.

T h e p h a r m a c e u t i c a l i n d u s t r y

FINDINGS

The pharmaceutical industry has been the

first to take advantage of the potentials of ap-

plied molecular genetics. Ult imately, i t wil l

probably benefit more than any other, with the

largest percentage of its products depending on

advances in genetic technologies. Already,

micro-organisms have been engineered to pro-duce human insulin, interferon, growth hor-mone, urokinase (for the treatment of bloodclots), thymosin-a 1 (for controlling the immuneresponse), and somatostatin (a brain hormone).(See figure 2.)

The products most likely to be affected bygenetic engineering in the next 10 to 20 yearsare non protein compou nd s like most antibiotics,and protein compounds such as enzymes andantibodies, and many hormones and vaccines.

Improvements can be made both in the prod-ucts and in the p rocesses by wh ich they are p ro-du ced. Process costs may be lowered an d evenentirely new prod ucts developed .

The most advanced applications today are inthe field of hormon es. While certain h ormoneshave already proved useful, the testing of others has been hindered by their scarcity andhigh cost. Of 48 human hormones that havebeen identified so far as possible candidates forproduction by genetically engineered micro-organisms, only 10 are used in current m edical

practice. The other 38 are not, partly becausethey have been available in such limited qu an-tities that tests of their therapeutic value havenot been possible.

Genetic technologies also open up new ap-proaches for vaccine developm ent for such in-tractable parasitic and viral diseases as amebicdysentery, trachom a, hepatitis, and malaria. Atpresent, the vaccine m ost likely to be p rodu cedis for foot-and-mouth disease in animals. How-ever, should any one of the vaccines for humandiseases become available, the social, economic,and political consequences of a decrease in mor-bidity and mortality would be significant. Manyof these diseases are particularly prevalent inless indu strialized coun tries; the developmentsof vaccines for them m ay p rofound ly affect thelives of tens of millions of people.



Ch. 1 Summary: Issues and Options q 5

Figure 1.— Recombinant DNA: The Technique of Recombining GenesFrom One Species With Those From Another

Electron micrograph of the DNA, which is the plasmid SPO1from Bacillus subtilis. This plasm id which has beensliced open is used for recombinant DNA research

Donor DNA

in this bacterial host

amount of DNA

SOURCE: Office of Technology Assessment.

Expressionproduces

protein

For some pharmaceutical products, biotech-nology will compete with chemical synthesisand extraction from human and animal organs.Assessing the relative worth of each methodmust be done on a case-by-case basis. But forother products, genetic engineering offers the

only method know n that can ensure a plentifulsupply; in some instances, it has no competition.

By making a pharmaceutical available, genet-ic engineering may have two types of effects:

q Drugs that already have medical promise

Photo credits; Professor F. A. Eiserling, UCLA Molecular Biology Institute

Electron micrograph of Bacillus subtilis in the process ofcell division. The twisted mass in the center of each

daughter cell is the genetic material, DNA

Restriction enzymes recognize certain sites along the DNA

and can chemically cut the DNA at those sites. This makesit possible to remove selected genes from donor DNA mole-

cules and insert them into plasmid DNA molecules to form

the recombinant DNA. This recombinant DNA can then be

cloned in its bacterial host and large amounts of a desired

protein can be produced.

will be available in ample amounts for clin-ical testing . Inter feron, for example, can betested for its efficacy in cancer and viraltherapy, and h um an growth hormon e canbe evaluated for its ability to heal woun ds.

q Other ph armacologically active su bstances

for which no apparent use now exists willbe available in sufficient qu antities and atlow enough cost to enable researchers toexplore new uses. As a result, the potentialfor totally new therapies exists. Regulatoryproteins, for examp le, wh ich are an entire

76-565 0 - 81 - 2



6 . Impacts of Applied Genetics—Micro-Organ/ims, Plants, and Animals

Figure 2.-The Product Development Process

2. Tissues

Micro-organisms such as E co/I

I 7. cutting 6. Plasmid

14. Process

development

scale-up

16. Industrial

The development process begins by obtaining DNA either through organic synthesis (1) or derived from biological sources such as tissues(2). The DNA obtained from one or both sources is tailored to form the basic “gene” (3) which contains the genetic information to “code” for adesired product, such as human interferon or human insulin. Control signals (4) containing instructions are added to this gene (5). Circular DNAmolecules called plasmids (6) are isolated from micro-organisms such as E. coIi; cut open (7) and spliced back(8) together with genes and con-trol signals to form “recombinant DNA” molecules. These molecules are then introduced into a host cell (9).

Each plasmid is copied many times in a cell (10). Each cell then translates the information contained in these plasmids into the desired prod-uct, a process called “expression” (11). Cells divide (12) and pass on to their offspring the same genetic information contained in the parentcell.

Fermentation of large populations of genetically engineered micro-organisms is first done in shaker flasks (13), and then in small fermenters

(14) to determine growth conditions, and eventually in larger fermentation tanks (15). Cel lular extract obtained from the fermentation process isthen separated, purified (16), and packaged (17) either for industrial use (18) or health care applications.Health care products are first tested in animal studies (19) to demonstrate a product’s pharmacological activity and safety, in the United

States, an investigational new drug application (20) is submitted to begin human clinical trials to establish safety and efficacy. Followingclinical testing (21), a new drug application (NDA) (22) is fried with the Food and Drug Administration (FDA). When the NDA has been reviewedand approved by the FDA the product maybe marketed in the United States (23).

SOURCE: Genentech, Inc.



Ch. 1—Summary: Issues and Options q 7

class of molecules that control gene activi-ty, are present in the body in only minutequantities. Now, for the first time, they canbe recognized, isolated, characterized, andproduced in quantity.

The mere availability of a pharmacologicallyactive substance d oes not ensure its adoption inmedical practice. Even if it is shown to havetherapeutic usefulness, it may not succeed inthe marketplace.

The difficulty in predicting the economic im-pact is exemp lified by inter feron. If it is foun d tobe broad ly effective against both viral diseasesand cancers, sales would be in the tens of bil-lions of dollars annually. If its clinical effec-tiveness is foun d to be only against one or tw oviruses, sales would be significantly lower.

At the very least, even if there are no im-mediate medical uses for compounds producedby genetic engineering, their indirect impact onmed ical research is assu red. For the first time,almost any biological phenomenon of medicalinterest can be explored at the cellular level.These molecules are valuable tools for under-standing the anatomy and functions of cells.The knowledge gained may lead to the develop-ment of new therapies or preventive measuresfor diseases.

T h e c h e m i c a l i n d u s t r y

FINDINGS

The chemical industry’s primary raw materi-

al, petroleum, is now in limited supply. Coal isone appealing alternative; another is biomass, a

renewable resource composed of plant and ani-

mal material.

Biomass has been transformed by fermenta-

tion into organic chemicals like citric acid, etha-

nol, and amino acids for decades. Other organic

chemicals such as acetone, butanol, and fumaric

acid were at one time made by fermentation un-

t i l chem ica l p roduc t ion m e thods , com binedwith cheap oil and gas, proved to be more eco-

nomical. In theory, most any industrial organic

chemical can be produced by a biological proc-

ess.

Commercial fermentation using genetically

engineered micro-organisms offers several ad-

vantages over current chemical productiontechniques.

The use of renewable resources: starches,sugars, cellulose, and oth er componen ts of biomass can serve as the raw material for

synthesizing organic chemicals. With prop-er agricultural management, biomass canassure a continuous renewable supply forthe industry.The use of physically milder conditions:chemical processes often require high tem-peratures and extreme pressures. Theseconditions are energy intensive and pose ahazard in case of acciden ts. Biological proc-esses operate under milder conditions,wh ich ar e compatible with living systems.One-step production methods: micro-orga-nisms can carry out several steps in a syn-

thetic process, eliminating th e need for in-termediate steps of separation and puri-fication.

Decreased pollution: because biological

processes are highly specific in the reac-tions they catalyze, they offer control overthe products formed and decrease undesir-able side-products. As a result, they pro-duce fewer pollutants that require manage-ment and disposal,

The impact of this technology w ill cut acrossthe entire spectrum of chemical groups: plastics

and resin materials, flavors and perfumes mate-rials, synthetic rubber, medicinal chemicals,pesticides, and the primary products from pe-troleum that serve as the raw materials for thesynthesis of organic chemicals. Nevertheless,the specific products that will be affected ineach group can only be chosen on a case-by-casebasis, with the applicability of genetics de-pend ing on a variety of factors. Crud e estimatesof the expected economic impacts are in the bil-lions of dollars per year for dozens of chemicalswithin 20 years.

IND USTRY AND MANPO WER IMPACTSAlthough genetic engineering will develop

new techniques for synthes iz ing many sub-

stances, the direct displacement of any current

industry seems doubtful. Genetic engineering

should be considered simply another industrial

too l . Indus t r i e s w i l l p robab ly use gene t i c



8 . Impacts of Applied Genetics—Micro-Organisms, Plants, and Animals

engineering to maintain their positions in theirrespective markets. This is already illustratedby the variety of companies in the pharmaceu-tical, chemical, and energy ind ustries that haveinvested in or contracted w ith genetic engineer-ing firms. Some large comp anies are already d e-

veloping inh ouse genetic engineering researchcapabilities.

Any predictions of the number of workersthat w ill be required in the prod uction phase of biotechnology will depend on the expectedvolum e of chemicals that will be produ ced. Atpresent, this figure is unknown. An estimated$15 billion worth of chemicals maybe manufac-tured by biological processes. This will employappr oximately 30,000 to 75,000 workers for su-pervision, services, and production. Whetherthis will represent a n et loss or gain in the n um -

ber of jobs is difficult to predict since new jobsin biotechnology will probably displace some of those in trad itional chemical produ ction.

Food proc e ss ing indus t ry

FINDINGS

Genetics in the food processing ind ustry canbe used in two w ays: to design micro-organismsthat transform inedible biomass into food forhuman consumption or into feed for animals;and to design organisms that aid in food proc-essing, either by acting directly on the food

itself or by providing materials which can beadded to food.

The use of genetics to d esign organisms w ithdesired properties for food processing is anestablished practice. Fermented foods andbeverages have been made by selected strainsof mutant organisms (e.g., yeasts) for centuries.Only recently, however, have molecular tech-nologies opened up new possibilities. In par-ticular, large-scale availability of enzymes willplay an increasing role in food processing.

.The applications of molecular genetics are

likely to appear in the food processing industryin piecemeal fashion:

q Inedib le biomass, human and an imalwastes, and even various industrial efflu-ents are now being transformed into edible

q

micro-organisms high in protein content(called single-cell protein or SCP). Its p res-ent cost of prod uction in the Un ited Statesis relatively high, and it mu st compete withcheaper sources of protein such as soy-beans and fishmeal, among others.

Isolated successes can be anticipated forthe production of such food additives asfructose (a sugar) and the synth etic sweet-ener aspartame, and for improvements inSCP production.

An industrywide impact is not expected in thenear future because of several major conflictingfactors:

q

q

q

The basic knowledge of the genetic charac-teristics that could improve food has notbeen adequ ately d eveloped.The food processing ind ustry is conserva-

tive in its expenditures for R&D to improveprocesses. Generally, only one-third to one-half as much is allocated for this pu rpose asin technologically intensive industries.Products made by new microbial sourcesmust satisfy the Food and Drug Adminis-

tration’s (FDA) safety regulations, which in-clude undergoing tests to prove lack of harmful effects. It may be possible toreduce the amount of required testing bytransferring the desired gene into micro-organisms that already meet FDA stand-ards.

The use o f gene t ica l ly engineered

mic ro - organ i sms in the e nv i ronme n t

FINDINGS

Genetically engineered micro-organisms are

being designed now to perform in three areas

(aside from agricultural uses) that require theirlarge-scale release into the environment:

. mineral leaching and recovery,

. enhanced oil recovery, and

q pollution control.

All of these are characterized by:

q the use of large volumes of micro-orga-

nisms,decreased control over the behavior and

fate of the micro-organisms,




the p ossibility of ecological disru ption, an dless development in basic R&D (and morespeculation-) than in the industries in whichmicro-organisms are used in a controlledenvironment.

MINERAL LEACHING AND RECOVERY

Bacteria have been used to leach metals, such

as uranium and copper, from low-grade ores.

Although there is reason to believe leaching

ability is under genetic control in these orga-

nisms, practically nothing is known about the

precise mechanisms involved. Therefore, the

application of genetic technologies in this arearemains speculative. Progress has been slow in

obtaining more information, part ly because

very little research has been conducted.

In addition to leaching, micro-organisms can

be used to recover valuable metals or eliminate

polluting m etals from dilute solutions such as in-dustrial waste streams. The process makes useof the ability of micro-organisms to bind metalsto their surfaces and then concentrate them in-ternally.

The economic competitiveness of biologicalmethod s is still unproved , but genetic mod ifica-tions have been attempted only recently. Thecost of producing the micro-organisms has beena m ajor consideration. If it can be redu ced, theapp roach might be useful.

ENHANCED OIL RECOVERY

Many methods have been tried in efforts torem ove o i l f rom the g round w hen na tu ra l

expulsive forces alone are no longer effective.

Injecting chemicals into a reservoir has, in many

cases, aided recovery by changing the oil’s flow

characteristics.

Micro-organisms can produce the necessary

chemicals that help to increase flow. Theoreti-

ca l ly , t hey can a l so be g row n in the w e l l s

themselves, producing those same chemicals in

situ. The currently favored chemical, xanthan,

is far from ideal for increasing flow. Genetic

engineering should be able to produce chem-icals with more useful characteristics.

The current research approach, funded by

the Dep artment of Energy (DOE) and ind epend-

ently by various oil companies, is a two-phase

process to find micro-organisms th at can func-tion in an oil reservoir environment, and then toimprove their characteristics genetically.

The genetic alteration of micro-organisms toproduce chemicals useful for enhanced oil re-covery has been mor e successful tha n the alter-ation of micro-organisms that may be used insitu. However, rDNA technology has not beenapplied to either case. All attempts have em-ployed artificially induced or naturally occur-ring mutations.

POLLUTION CONTROL

Many micro-organisms can consume various

kind s of pollutants, changing them in to relative-

ly harmless materials before they die. These

micro-organisms always have had a role in

“natural” pollution control: nevertheless, cities

have resisted adding microbes to their sewerage

systems. Although the Environmental Protec-

tion Agency (EPA) has not recommended addi-

tion of bacteria to municipal sewerage systems,

it suggests that they might be useful in smaller

installations and for specific problems in large

systems. In major marine spills, the bacteria,

yeast, and fungi already present in the water

part icipate in degradation. The usefulness of

added microbes has not been demonstrated.

Nevertheless, in 1978, the estimated market

of biological products for pollution control was$2 million to $4 million per year, divided among

some 20 companies; the potential market wasestimated to be as much as $20 million per year.

To date, genetically engineered strains have

not b een app lied to pollution p roblems. Restrict-

ing factors include the problems of liability in

the event of health, economic, or environmental

damage; the contention that added organisms

are not likely to be a significant improvement;

and the assumption that selling microbes rather

than products or processes is not likely to beprofitable.

Convincing evidence that microbes could re-move or degrade an intractable pollutant would

encourage their application. In the meantime,

however, these restrictions have acted to inhibit

the research necessary to produce marked im-

provements.



10 . Impacts of Applied Genetics —Micro-Organisms, Plants, and Animals

CONSTRAINTS IN USING GENETIC ENGIN EERINGTECHNOLOGIES IN OPEN ENVIRONMENTS

The genetic data base for the p otentially use-ful micro-organisms is lacking. Only the sim-plest methods of mutation and selection for de-sirable properties have been used thus far.These are the only avenues for improvement

until more is learned about the genetic mech-anisms.

Even w hen the scientific know ledge is avail-able, two other obstacles to the use of geneti-

systems on a scale large enough to exploit theirbiological activity. This will necessitate a con-tinual dialog among microbial geneticists, geolo-gists, chemists, and engineers; an interdisci-plinary app roach is required that recognizes theneeds and limitations of each discipline.

The second obstacle is ecological. Introducinglarge numbers of genetically engineered micro-organisms into the environment might lead toecological d isrupt ion or detrim ental effects onhuman health, and raise questions of legal lia-

cally engineeredThe first is the

micro-organisms will remain. bility.—

need to develop engineered

I s s u e an d O p t io n s — Bio tech n o lo g y

I S S U E: H ow c a n the F e de r a l G ove r n -ment promote advances in bio-

t e c h n o l o g y a n d g e n e t i c e n g i -n e e r i n g ?

The United States is a leader in applyinggenetic engineering and biotechnology to in-du stry. One reason is the long-standing commit-ment by the Federal Government to the fundingof basic biological research; several decades of support for some of the most esoteric basicresearch has unexpectedly provided the foun-dation for a highly useful technology. A secondis the availability of venture capital, wh ich h asallowed the formation of sma ll, innovative com-

panies that can bu ild on the basic research.The chief argument for Governm ent subsidi-

zation for R&D in biotechnology and geneticengineering is that Federal help is needed inareas such as general (generic) research or high-ly sp eculative investigations not now being de-veloped by indu stry. The argument against th eneed for this supp ort is that ind ustry w ill devel-op everything of commercial value on its ow n.

A look at what ind ustry is now attempting in-dicates that sufficient investment capital isavailable to pu rsue sp ecific man ufacturing ob-

jectives. Some high-risk areas, however, thatmight be of interest to society, such as pollutioncontrol, may justify pr omotion by th e Govern-ment, w hile other, such as enh anced oil recov-ery might m ight not be profitable soon enoughto attract investment by ind ustry.

OPTIONS:

A. Congress could allocate funds specifically for genetic engineering and biotechnology R&Din the budget of appropriate agencies.

Congress could promote two types of pro-grams in biotechnology: those with long-rangepayoffs (basic research), and those that industryis not willing to und ertake but that migh t be inthe national interest.

B. Congress could establish a separate Institute of Biotechnology as a funding agency.

The merits of a separate institute lie in thepossibility of coordinating a wide range of ef-forts, all related to biotechnology. On the otherhand, biotechnology and genetic engineeringcover such a broad range of disciplines that anew funding agency would overlap the man-dates of existing agencies. Furthermore, thecreation of yet another agency carries with it allthe disadvantages of increased bureaucracy andcompetition for funds at the agency level.

C. Congress could establish research centers inuniversities to foster interdisciplinary ap-

proaches to biotechnology. In addition, a program of grants could be offered to train scien-

tists in biological engineering.

The successful use of biological techniques inindustry depends on a multidisciplinary ap-proach involving biochemists, geneticists, mi-crobiologists, process engineers, and chemists.



Ch. 1—Summary: lssues and Options q 11

Little is now being done publicly or privately todevelop the expertise necessary.

D. Congress could use tax incentives to stimulate biotechnology.

The tax laws could be used to stimulate bio-technology by expanding the supply of capitalfor small, high-risk firms, which are generallyconsidered more innovative than establishedfirms because of their willingness to undertakethe risks of innovation. In ad dition to focusingon the supply of capital, tax policy could at-tempt to directly increase the profitability of potential growth compan ies.

A tax incentive could also be directed’ at in-creasing R&D expenditures. It has been sug-gested that companies be permitted to take taxcredits: 1) on a certa in percentage of their R&Dexpenses; and 2) on contributions to universities

for research.

E. Congress could improve the conditions underwhich U.S. companies collaborate with aca-

demic scientists and make use of the technology developed in universities, which has beenwholly or partly supported by tax funds.

Developments in genetic engineering havekindled interest in this option. Under legislationthat has recently passed both Houses of Con-

gress, small businesses and universities may re-tain title to inventions developed un der federal-ly funded research. Currently, some Federalagencies award contractors these exclusiverights, while others insist on the nonexclusivelicensing of inven tions.

F. Congress could mandate support for specific research tasks such as pollution control using microbes.

Microbes may be u seful in d egrading int rac-table wastes and pollutants. Current research,however, is limited to isolating organisms fromnatural sources or from mutated cultures. Moreelaborate efforts, involving rDNA techniques orother form s of microbial genetic exchan ge, willrequire add itional fund ing.

G. Most efforts could be left to industry and each

Government agency allowed to develop programs in the fields of genetic engineering and biotechnology as it sees fit.

Generic research w ill probably n ot be under-taken by any one company. Leaving all R&D inindustry’s hands w ould still prod uce major com-mercial successes, but does not ensure the de-velopment of needed basic general knowledgeor the undertaking of high-risk projects.

A g r i c u l t u r eThe complexity of plants and animals pre-

sents a greater challenge to adv ances in app liedgenetics than that posed by micro-organisms.Nevertheless, the su ccessful genetic manipu la-tion of microbes has encouraged researchers inthe agr icultu ral sciences. The new tools will beused to complement, but not replace, the well-established practices of plant and animalbreeding.

T h e app l i c a t ions o f gene t i c s to p lan t s

FINDINGS

It is impossible to exactly determine the ex-

tent to which applied genetics has directly con-

tributed to increases in plant yield because of

s imul taneous improvements in farm manage-

ment, pest control, and cropping techniquesusing herbicides, irrigation, and fertilizers.Nevertheless, the impacts of breeding technol-ogies have been extensive.

The plant breeder’s approach is determinedfor the most part by the particular biologicalfactors of the crop being bred . The new genetictechnologies potentially offer additional tools toallow development of new varieties and evenspecies of plants by circumven ting current bio-logical barriers to the exchange of geneticmaterial.

Technologies developed for classical plantbreeding and those of the new genetics shouldnot be viewed as being competitive; they areboth tools for effectively man ipu lating genetic



12 . Impacts of Applied Genetics— Micro-Organisms, Plants, and Animals

information. One n ew technology -e.g., proto-plasm fusion, or the a rtificial fusion of two cells–allows breeders to overcome incompatibilitybetween plants. But the plant that may resultstill must be selected, regenerated, and eval-uated und er field conditions to ensure that thegenetic change is stable and that the attributesof the new variety meet commercial require-ments.

In theory, the new technologies will expan dthe capability of breeders to exchange geneticinformation by overcoming natural breedingbarriers. To date, however, they have not had awidespread impact on the agricultural industry.

As a note of caution, it mu st be emph asizedthat no plant can possess every desirable trait.There will always have to be some tradeoff;

often quality for quantity, such as increasedprotein content but d ecreased yield.

NEW GENETIC TECHNO LOGIES FORPLANT BREEDING

The new technologies fall into two categories:those involving genetic transformations

through cell fusion and those involving the in-sertion or mod ification of genetic informationthrough the cloning of DNA and its vectors.Techniques are available for manipulatingorgans, tissues, cells, or p rotoplasts in culture;for regenerating plants; and for testing thegenetic basis of novel traits. So far these tech-niques are r outine only in a few species.

The approach to exploiting molecular biologyfor plant breeding is similar in some respects tothe genetic manipulation of micro-organisms.However, there is one major conceptual dif-

A plantlet of Ioblolly pine grown in Weyerhaeuser Co.’stissue culture laboratory. The next step in this procedure

is to transfer the plantlet from its sterile and humidenvironment to the soil

Photo credits: Weyerhaeuser Co.

A young Douglas fir tree propagated 4 years ago from asmall piece of seedling leaf tissue. Three years ago this wasat the test-tube stage seen in the Ioblolly pine photograph



Ch. 1—Summary: Issues and Options . 13

ference. In micro-organisms, the changes made

on the ce l lu l a r l eve l a r e the goa l s o f t he

manipulation. With crops, changes made on the

cellular level are meaningless unless they can bereproduced in the entire plant as well. There-fore, unless single cells in culture can be

selected and grown into mature plants and thedesired traits expressed in the mature plant–

procedures which at this time have had limited

success—the b enefits of genetic engineering w illnot be widely felt in plant b reeding.

M odera te success has been ach ieved fo rgrowing cells in t issue culture into mature

plants. Tissue culture programs of commercial

significance in the United States include theasparagu s, citrus fruits, pineap ples, and straw-berries. Breeders have had little success, how-ever, in regenerating mature plants of wideagronomic imp ortance) such as corn and w heat.

Some success can be claimed for engineeringchanges to alter genetic makeup. Both the stableintegration of genetic material into a cell andthe fusion of genetically different cells are stilllargely experimental techniques. Technicalbreakthroughs have come on a species-by-species basis, but key discoveries are not oftenapplicable to all plants. Initial results suggestthat agronomically important traits, such asdisease resistance, can be transferred from onespecies to another. Limited success has alsobeen shown in attempts to create totally new

species by fusing cells from different genera.Attempts to find both suitable vectors and genesfor transferring one plant genes to another areonly now beginning to show promise.

CONSTRAINTS ON USING MOLECULARGENETICS FOR PLANT IMPROVEMENTS

Molecular engineering has been impeded by alack of answ ers to basic questions in molecularbiology and plant p hysiology owing to insuffi-cient research. Federal funding for plant molec-ular genetics in agriculture h as come primarilyfrom the U.S. Department of Agriculture

(USDA) and the National Science Foundation(NSF). In USDA, research support is channeledprimarily through the flexible competitivegrants prog ram (fiscal year 1980 budget of $15million) for the sup port of new research d irec-tions in plant biology. The total support for the

plant sciences from NSF is approximately $25million, only $1 million of w hich is sp ecificallydesignated for plant gen etics.

The shortage of a trained workforce is asignificant constraint. Only a few universitieshave expertise in both p lants and molecular bi-ology. In ad dition, there are only a few peoplewho have the ability to work with modern mo-lecular techniques related to whole plant prob-lems. As a result, a business firm could easilydevelop a capability in this area exceeding thatat any individual U.S. university. However, thebuilding of industrial laboratories and subse-quen t hiring from th e un iversities could easilydeplete the expertise at the university level.With the recent investment activity by manybioengineering firms, this trend has alreadybegun; in the long-run it could hav e serious con-sequences for the quality and quantity of uni-versity research.

GENETIC VARIABILITY, CROP VULNERABILITY,AND THE STORAGE OF GERMPLASM

Successful plant breeding is based on theavailability of genetically diverse plants for theinsertion of new genes into plants. The numberof these plants has been d iminishing for a varie-ty of reasons. However, the rate and extent of this trend is unknown; the data simply do notexist. Therefore, it is essential to have an ade-quate scientific understanding of how much ge-netic loss has taken place and how much germ-plasm (the total genetic variability available to aspecies) is needed. Neither of these questionscan be answ ered comp letely at this time.

Even if-genetic needs can be ad equately iden-tified, there is disagreement abou t the quantityof germplasm to collect. Furthermore, the ex-tent to which the new genetic technologies willaffect genetic variability, vulnerability, or thestorage technologies of germplasm has not beendeterm ined. As a result, it is currently d ifficult,if not impossible, to state how much effortshould be expended by the N ational Germplasm

System to collect, maintain, and test new generesources (in this case as seed).

Finally, even if an adequate level of geneticvariability can be assessed, the real p roblem of vulnerability—the practice of planting only a



Ch. 1—Summary: Issues adoptions q 1 5

Advances in reproduc t ive b io logy and

the i r e f f e c t s on an ima l improv e me n t

FINDINGS

Much improvement can be made in the germ-plasm of all major farm animal species using ex-

isting technology. The expanded use of artificialinsemination (AI) with stored frozen sperm, es-pecially in beef cattle, wou ld benefit both pro-du cers and consumers. New techniques for syn-chronizing estrus should encourage the wideruse of AI. Various manipulations of embryos

wil l f ind l imi ted use in producing breeding

stocks, and sex selection and twinning tech-

niques should be available for limited applica-

tions w ithin the n ext 10 to 20 years.

The most important technology in reproduc-

tive physiology will continue to be AI. Due in

part to genetic improvement, the average milk yield of cows in the United States has more than

doubled in the past 30 years, while the totalnumber of milk cows has been reduced by more

than h alf. AI along with improved managementand the availability and use of accurate p rogenyrecords on breeding stock have caused thisgreat increase. (See figure 3.)

The improvement lags behind w hat is theo-retically possible. In practice, the observed in-crease is about 100 lb of milk p er cow p er year,wh ile a hyp othetical breeding program using AIwou ld resu lt in a yearly gain of 220 lb of milk per cow. The biological limits to this rate of gainare not known.

In comparison with dairy cattle, the beef cat-tle ind ustry h as not ap plied AI technology wide-ly. Only 3 to 5 percent of U.S. beef is art ificiallyinseminated, compared to 60 percent of theda iry herd . This low ra te for beef cattle can beexplained by several factors, including man age-ment techniques (range v. confined housing)and the conflicting objectives of individualbreeders, ranchers, breed associations, andcommercial farmers.

The national calf crop—calves alive at wean-ing as a fraction of the total number of cows ex-posed to breeding each year—is only 65 to 81percent. An improvemen t of only a few p ercent-age points through AI would r esult in savings of

hu nd reds of millions of dollars to produ cers andconsumers.

Coupled with a technology for estrus-cycleregulation, the u se of AI could be expand ed forboth d airy and beef breeding. Embryo transfer

technology, already well-developed but stillcostly, can be used to p roduce valuable breed-ing stock. Sexing technology, which is not yetperfected, wou ld be of enorm ous benefit to thebeef industry because bulls grow faster thanheifers.

In the case of animals other than cows:

q

q

q

q

q

Expanded use of AI for swine productionwill be encouraged by the strong trend toconfinement housing, although the poorability of boar sperm to w ithstand freezingwill continue to be a handicap.

The benefits of applied genetics have notbeen realized in sheep production becauseneither AI nor performance test ing has

been used. As long as the use of AI con-tinues to be limited by the inability tofreeze semen an d by a lack of agents on themarket for synchronizing estrus, no rap idmajor gains can be expected.Increasing interest in goats in the UnitedStates and the deman d for goat prod uctsthroughout the world, should encourageattention to the genetic gains that the useof AI and other technologies make possible.

Poultry breeders will continue to concen-trate on improved egg production, growthrate, feed efficiency, and redu ced body fatand diseases. The use of frozen semenshould increase as will the use of AI anddwarf broiler breeders.Genetics applied to production of fish,mollusks, and crustaceans in either naturalenvironments or manmade culture systemsis only at the rudimentary stage.

Breeders must have reliable informationabout th e genetic value of the germplasm th ey

are considering introducing. Since farmers donot have the resources to collect and processda ta on the performance of animals other thanthose in their own herds, they must turn to out-side sources. The National Cooperative DairyHerd Improvement Program (NCDHIP) is a mod-



Ch. 1—Summary: Issues and Options . 17

el information system and could be adapted toother species.

Selection—deciding which animals to mate—is the breeder’s most basic tool. When goingoutside his herd to purchase new germplasm,the breeder needs impartial information aboutthe qu ality of the available germplasm. N CDHIPhad recorded 2.8 million of th e 10.8 million U.S.

dairy cattle in 1979. In 1978, cows enrolled in

the official plans of NCDHIP outproduced cows

not enrolled by 5,000 lb of milk per cow, repre-

sentin g 52 percent more m ilk per lactation.

No comparable information system exists forother types of livestock. Beef bulls, for example,continue to be sold to a large extent on the basisof pedigrees, but with relatively little objectiveinformation on their genetic merit. Data ondairy goats in the United States became avail-

able through NCDH IP for the first time in late1980. No nationwide information systems exist

for other species, although pork production inthe United States wou ld greatly benefit from anational swine testing p rogram.

The more esoteric methods of genetic manip-ulation will probably have little impact on the

production of animals or animal products with-in the next 10 years. other in vitro manipula-tions, such as cloning, cell fusion, the produc-tion of chimeras, and the use of rDNA tech-niques, will continue to be of intense interest,especially for research purposes. It is less likely,however, that they w ill have w idespread p rac-tical effects on farm production in this century.

Each technique requires more research andrefinement. Until specific genes of farm animalscan be identified and located, no direct genemanipulation will be practicable. In addition

this will be difficult because most traits of im-portance are du e to multiple genes.

Issue a n d O p t i o n s — A n i m a l s

ISSUE: H o w ca n t h e F ed e r a l Go v er n -ment improve the germplasm of major farm animal species?

OPTIONS:

A. Programs like the NCDHIP could have in-

creased governmental participation and fund-ing. The efforts of the Beef Cattle Improve- ment Federation to standardize procedures could receive active support, and a similar information system for swine could be estab-lished.

The fastest and least expensive way to up-grade breeding stock in the United States isthrough effective use of information. Computertechnology, along with a network of local repre-

* *

The wide variety of applications for genetic

engineering is summarized in figure 4. Geneticscan be used to improve or increase the qualityand outpu t of plants and animals for direct useby man. Alternatively, materials can be ex-tracted from plants and animals for use in food,chemical, and pha rmaceutical ind ustries.

sentatives for data collecting, can provide theindividual farmer or breeder with accurate in-formation on the available germp lasm so that hecan make h is own breeding d ecisions.

This option implies that the Federal Govern-ment w ould p lay such a role in new programs,

and expand its role in existing ones.

B. Federal funding could be increased for basic research in total animal improvement.

This option, in contrast to op tion A, assumesthat it is necessary to maintain or expand basicR&D to generate new knowledge that can beapp lied to th e produ ction of improved animalsand animal products.

*

Biological materials can also be converted to

useful products. In this process, genetic engi-neering can be used to d evelop m icro-organismsthat will carry out the conversions. Therefore,genetic manipulation cannot only provide moreor better biological raw materials but can alsoaid in th eir conversion to useful prod ucts.



18 . The Impacts of Genetics: Applications to Micro-Organisms, Plants, and Animals

Figure 4.—AppIications of Genetics

IAGRICULTURAL

INDUSTRY I

Plants

Animals

(lncrease/Improve, Output)


I n s t i t u t i o n s a n d s o c i e t y

Regula t ion o f gene t ic engineer ing

FINDINGS

No evidence exists that any unexpectedlyharmful genetically engineered organism hasbeen created. Yet few experts believe that mo-lecular genetic techniques are totally withoutrisk to health and the environment. Informationthat has proved useful in assessing the risksfrom these techniques has come from threesources: experiments designed specifically totest the consequences of working with rDNA,experiments designed for other purposes but

relevant to rDNA, and scientific meetings andworkshops.

A p rogram of risk assessment w as establishedat N IH in 1979 to cond uct experiments and col-late relevant information. It assesses one formof genetic engineering, rDNA. On the basis of these data, conjectured, inadvertent risk is

generally regarded as less likely today thanoriginally susp ected. Risk du e to the man ipu la-tion of genes from organ isms know n to be haz-ardous is considered to be more realistic. There-fore, microbiological safety precautions that are



Ch. 1—Summary: lssues and Options . 19

appropriate to the use of the micro-organismsserving as the source of DNA ar e required. Nev-ertheless, it has not been demonstrated thatcombining those genes in the form of rDNA isany more hazardous than the original source of the DNA.

Perceptions of the nature, magnitud e, and ac-ceptability of the risk differ. In addition, publicconcern has been expressed about possiblelong-range impacts of genetic engineering. Inthis context, the problem facing the policy-maker is how to add ress the risk in a w ay thataccommodates the perceptions and values of those who bear it.

The NIH Guidelines for Research InvolvingRecombinant DNA Molecules and existing Fed-eral laws appear adequate in most cases to dealwith the risks to health and the environmentpresented by genetic engineering. However, the

Guidelines are not legally binding on ind ustry,and no single statute or combination will clearlycover all foreseeable commercial applications of genetic engineering.

The Guidelines are a flexible evolving over-sight mechanism that combines technical exper-tise with public participation. They cover themost widely used and possibly risky moleculargenetic technique–rDNA–prohibiting experi-ments u sing d angerous toxins or pathogens andsetting containment standards for other poten-tially hazard ous experiments. Although compli-

ance is mandatory only for those receiving NIHfunds, other Federal agencies follow them, andindustry has proclaimed voluntary compliance.Rare cases of noncomp liance have occurr ed inuniversities but have not posed risks to healthor the environmen t. As scientists have learnedmore about rDNA and molecular genetics, therestrictions have been progressively and sub-stantially relaxed to the point where 85 percentof the experiments can now be done at thelowest containment levels, and virtually allmonitoring for compliance now rests with ap-proximately 200 local self-regulatory commit-

tees called institutional biosafety committees(IRCs). (See table 1.)

Und er the Guidelines, NIH serves an impor-tant oversight role by sponsoring risk assess-

Table I.—Containment Recommended by theNational institutes of Health

Biological—Any combination of vector and host must bechosen to minimize both the survival of the systemoutside of the laboratory and the transmission of thevector to nonlaboratory hosts. There are three levelsof biological containment:

HVl– Requires the use of Escherichia coli K12 orother weakened strains of micro-organisms thatare less able to live outside the laboratory.

HV2– Requires the use of specially engineered strainsthat are especially sensitive to ultraviolet light,detergents, and the absence of certainuncommon chemical compounds.

HV3– No organism has yet been developed that canqualify as HV3.

Physical—Special laboratories (P1-P4)

Pl— Good laboratory procedures, trained personnel,wastes decontaminated

P2– Biohazards sign, no public access, autoclave inbuilding, hand-washing facility

P3– Negative pressure, filters in vacuum line, class IIsafety cabinets

P4– Monolithic construction, air locks, all air

decontaminated, autoclave in room, allexperiments in class Ill safety cabinets (glovebox), shower room


ment programs, certifying new host-vector sys-tems, serving as an information clearinghou se,and coordinating Federal and local activities.Limitations in N IH’s oversigh t are tha t: it lackslegal authority over indu stry; its procedu res foradvising industry on large-scale projects havenot incorporated sufficient expertise on large-scale fermentation technology; its monitoring

for either comp liance or consistent app licationof the Guidelines by individuals or institutions isvirtually nonexistent; and it has not systemati-cally evaluated other techniques, such as cell fu-sion, that might present risks.

Federal laws on health and environment willcover most comm ercial applications of geneticengineering. Produ cts such as dru gs, chemicals,and foods can be regulated by existing laws.However, uncertainty exists about the regula-tion of either production methods using engi-neered micro-organisms or their intentionalrelease into the environment, when the risk hasnot been clearly demonstrated. While a broadinterpretation of certain statutes, such as theOccupational Safety and Health Act and theToxic Substances Control Act, migh t cover these




situations, regulatory actions based on such in- substantial regulatory authority over commer-terpretations could be challenged in court. In cial genetic engineering have not yet officially

any evident those agencies that could have acted to assert that au thority.

I s s u e an d O p t io n s — Reg u la t i o n

ISSUE: How could Congress address th erisks presented by genetic engi-neer ing?

OPTIONS:

A. Congress could maintain the status quo by let- ting NIH and the regulatory agencies set the Federal policy.

Congress might determine that legislation toremedy the limitations in current Federal over-sight w ould result in un necessary and burd en-some regulation. No kn own harm to h ealth orthe environment has occurred under currentregulation. Also the agencies generally have thelegal authority and expertise to adapt to mostnew problems posed by genetic engineering.

The disadvantages are the lack of a central-ized, uniform Federal response to the problem,and the possibility that risks associated withcommercial applications will not be adequ atelyadd ressed. Conflicting or red und ant regulationsof different agencies would result in unneces-sary burd ens on those regulated.

B. Congress could require that the Federal Inter- agency Advisory Committee on Recombinant

DNA Research prepare a comprehensive report on its members’ collective authority to regulate rDNA and on their regulatory intentions.

The Industrial Practices Subcommittee of thisCommittee has been studying agency authorityover commercial rDNA activities. Presently,there is little official guidance on regulatory re-quirements for companies that may soon mar-ket produ cts made by rDN A methods. A con-gressionally mandated report would ensure fullconsideration of these issues by the agenciesand expedite the process. On the other hand,

the agencies are studying the situation, whichmust be done before they can act. Also, it isoften easier and more efficient to act on eachcase as it arises, rather than on a hypotheticalbasis before the fact.

C. Congress could require that all recombinantDNA activity be monitored for a limited num-

ber o f years.

This represents a “wait and see” position by

Congress and the middle ground between thestatus quo and full regulation. It recognizes and

balances the following factors: 1) the absence of

demonstrated harm to human health or the en-

vironment from genetic engineering; 2) the con-

tinuing concern that genetic engineering pre-sents risks; 3) the lack of sufficient knowledge

and experience from which to make a final judg-

ment; 4) the existence of an oversight mech-anism that seems to be working well, but thathas clear limitations with respect to commercial

activities; 5) the virtual abolition of Federal

monitoring of rDNA activities by recent amend-

ments to the Guidelines; and 6) the expected in-crease in commercial genetic engineering.

This option would provide a data base that

could be used for: 1) determining the effec-

t iveness o f vo lun ta ry com pl i ance w i th the

Guidelines by industry, and mandatory compli-

ance by Federal grantees; 2) determining thequality and consistency of the local self-regu-

latory actions; 3) continu ing a formal risk assess-ment program; 4) identifying vague or conflict-

ing provisions of the Guidelines for revision; 5)

identifying emerging trends or p roblems; and 6)

tracing any long-term adverse impacts on health

or the environment to their sources.

The obvious disadvantage of this optionwould be the required paperwork and effort byscientists, universities, corporations, and theFederal Government.

D. Congress could make the NIH Guidelines ap-

plicable to all rDNA work done in the United States.

This option would eliminate any concernabout the effectiveness of voluntary compliancewith the Guid elines, and it has the ad vantage of



Ch. 1—Summary: Issues adoptions q 21

using an existing oversight mechanism. The major changes that wou ld have to be mad e in thearea of enforcement. Present p enalties for non-compliance—susp ension or termination of re-search funds-are obviously inapplicable to in-du stry. In add ition, procedu res for monitoring

compliance would have to be strengthened.

The main disadvantage of this option is thatNIH is not a regu latory agency. Since NIH hastraditionally viewed its mission as promotingbiomedical research, it would have a conflict of interest between regulation and promotion.One of the r egulatory agencies could be giventhe authority to enforce the Guidelines.

E. Congress could require an environmental impact statement and agency approval before any genetically engineered organism is inten-

tionally released into the environment.

There have been numerous cases where ananimal or plant sp ecies has been introduced intoa new environment and has spread in an un con-trolled and undesirable fashion. Yet in pollutioncontrol, mineral leaching, and enhanced oilrecovery, it m ight be desirable to release largenumbers of engineered micro-organisms intothe environment.

The Guidelines curren tly proh ibit deliberaterelease of any organism containing rDNA with-out ap proval of NIH. One disadvan tage of thisprohibition is that it lacks the force of law.Another is that app roval may be granted on afind ing that the release wou ld pr esent “no sig-nificant risk to health or the environment;” atougher or more specific standard may be de-sirable.

A required study of the possible conse-quences of releasing a genetically engineeredorganism would be an important step in ensur-ing safety. An impact statement could be filedbefore permission is granted to release theorganism. However, companies and individuals

might be discouraged from developing usefulorganisms if this process became too burden-some and costly.

F. Congress could pass legislation regulating all types and phases of genetic engineering from research through commercial production.

This option would deal comprehensively anddirectly with the risks of novel moleculargenetic techniques. A specific statute wouldeliminate the uncertainties over the extent towhich present law covers particular applica-tions of genetic engineering and any concerns

about the effectiveness of voluntary compliancewith the Gu idelines. Alternatively, the legisla-tion could take the form of amending existinglaws to clarify their applicability to geneticengineering.

Other molecular genetic techniques, whilenot as w idely used and effective as rDNA, raisesimilar concerns. Of the current techniques, cellfusion is the prime cand idate for being treatedlike rDNA in any regu latory framew ork. No risk assessment of this technique has been done, andno Federal oversight exists.

The principal argument against this option isthat the current system ap pears to be workingfairly well, and the limited risks of the tech-niques may not warrant the significantly in-creased regulatory burden that would resultfrom such legislation.

G. Congress could require NIH to rescind theGuidelines.

Deregulation would hav e the advantage of al-lowing money an d p ersonnel curren tly involvedin implementing the Guidelines at the Federaland local levels to be used for other pu rposes.

There are several reasons for retaining theGuidelines. Sufficient scientific concern existsfor the Guidelines to prohibit certain experi-ments and to require containment for others.Most experiments can be done at the lowest,least burdensome containment levels. NIH isserving an important role as a centralized over-sight and information coordinating body, andthe system h as been flexible enough in the p astto liberalize the restrictions as evidence in-dicated lower risk than originally thought.

H. Congress could consider the need for regulat-ing work with all hazardous micro-organisms

and viruses, whether or not they are genet-ically engineered.

It was not within th e scope of this study to ex-amine this issue, but it is an emerging one thatCongress may wish to consider.

76-565 0 - 81 - 3




P a ten t i n g li v i n g or g a n i s ms

On Jun e 16, 1980, in a 5-to-4 decision, the Su-preme Court ruled that a hum an-made micro-organism w as patentable under Federal patentstatutes. The decision wh ile hailed by some asassuring th is country’s technological futu re w as

at the same time denounced by others as creat-ing Aldous Huxley’s Brave New World. It will doneither.

FINDINGS

1. Meaning and Scope of the Decision.—Thedecision held tha t a patent could not be deniedon a genetically engineered micro-organism thatotherwise met the legal requirements for pat-entability solely because it was alive. It wasbased on the Court’s interpretation of a provi-sion of the patent law wh ich states that a patentmay be granted on “. . . any new and useful . . .

man ufacture, or composition of matter. . . .“ (35U.S.c. $101)

It is uncertain wh ether the case will serve asa legal precedent for patenting more complexorganisms. Such org anisms, however, w ill prob-ably not meet other legal prerequisites to paten-tability that w ere not at issue here. In any event,fears that the case would be legal precedentsometime in the distant future for patenting hu-man beings are unfounded because the 13thamendment to the Constitution absolutely pro-hibits own ership of hu mans.

2. Impact on the Biotechnology Industry.—Thedecision is not crucial to the d evelopment of theindu stry. It will stimulate innovation by encour-aging the dissemination of technical informa-tion that otherwise would have been main-tained as trade secrets because patents are pub-lic docum ents that fully describe the inven tions.In addition, the ability to patent genetically engi-neered micro-organisms will reduce the risksand uncertainties facing ind ividual compan iesin the commercial development of those orga-nisms and their prod ucts, but only to a limiteddegree because reasonably effective alterna-

tives exist. These are: 1) maintaining the orga-

nisms as trade secrets; 2) patenting microbio-logical processes and their pr odu cts; and 3) pat-enting inanimate components of micro-orga-nisms, such as genetically engineered p lasmids.

3. Impact on the Patent Law and the Patent and Trademark Office.— Because of the com plexity,

reproducibility, and mutability of living orga-nisms, the decision may cause some problemsfor a body of law designed m ore for inan imateobjects than for living organisms. It raises ques-tions about the p roper interpretation and appli-cation of the patent law requirements of novel-ty, nonobviousness, and enablement. In addi-tion, it raises questions about how broad thescope of patent coverage on important micro-organisms should be, and about the continuingneed for two statutes, the Plant Patent Act of 1930 and the Plant Variety Protection Act of 1970. These uncertainties could result in in-

creased litigation, making it more difficult andcostly for owners of p atents on living organismsto enforce their rights.

The impact on the Patent and Trademark Of-fice is not expected to be significant in the n extfew years. Although the nu mber of patent ap -plications on micro-organisms have almostdoubled during 1980, the approximately 200pending applications represent less than 0.2percent of those processed each year by the Of-fice. While the n um ber of such app lications isexpected to increase in the next few yearsbecause of of the decision and developments inthe field, the Office should be able to ac-commodate the increase. A few additional ex-aminers may be needed. ,

4. Impact on Academic Research.—Because thedecision m ay encourage academ ic scientists tocommercialize the results of their research, itmay inhibit the free exchange of information,but only if scientists rely on trade secrecyrather than patents to p rotect their inventionsfrom competitors in the marketplace. In this re-spect, it is not clear how molecular biology dif-fers from other r esearch fields w ith commercial

potential.




I ssue and Opt ions — P a t e n t i n g L i v i n g O r g a n i s m s

ISSUE: To wha t ex ten t could Congressprovide for or prohibi t the pat-ent ing of living organisms?

OPTIONS:

The Supreme Court stated that it was under-taking only the narrow task of determiningwh ether or not Congress, in enacting the patentstatutes, had intended a manmade micro-orga-nism to be excluded from patentability solelybecause it was alive. Moreover, the opinionspecifically invited Congress to overrule thedecision if it disagreed with the Court’s inter-pretation. Congress can act to resolve the ques-tions left unanswered by the Court, overrulethe d ecision, or d evelop a comprehensive statu-tory approach. Most importantly, Congress can

draw lines; it can decide which organisms, if any, should be p atentable.

A. Congress could maintain the status quo.

Congress could choose not to address theissue of patentability and allow the law to bedeveloped by the courts. The advantage of thisoption is that issues will be addressed as theyarise, in the context of a tangible, nonhypo-thetical case.

There are two d isadvantages to this option: auniform body of law may take time to develop;

and the Federal jud iciary is not designed to takesufficient account of the broader political andsocial interests involved.

B. Congress could pass legislation dealing with the specific legal issues raised by the Court's decision.

Many of the legal questions are so broad andvaried that they do not readily lend themselvesto statutor y resolution, The precise meaning of the requiremen ts for novelty, nonobviousness,and enablement as app lied to biological inven-tions will be most readily developed on a case-

by-case basis by the Patent Office and theFederal courts. On the other hand, some ques-tions are fairly narrow and well-defined; thus,they could be better resolved by statute. Themost important question is whether there is acontinuing need for the two plant protection

acts that grant ownership rights to plantbreeders who develop new and dist inctvarieties of plants.

C. Congress could mandate a study of the Plant

Patent Act of 1930 and the Plant Variety Pro- tection Act of 1970.

These Acts could serve as m odels for stud yingthe broader, long-term potential impacts of patenting living organisms. Such a stu dy w ouldbe timely not only because of the Cour t’s deci-sion, but also because of allegations that theActs have encouraged the planting of uniformvarieties, loss of genetic diversity, and increasedconcentration in the plant breed ing indu stry.

D. Congress could prohibit patents either on anyliving organism or on organisms other than

those already subject to the plant protection Acts.

By prohibiting patents on any living or-ganisms, Congress would be accepting thearguments of those who consider ownershiprights in living organisms to be immoral, or w hoare concerned about other potentially adverseimpacts of such patents, A total prohibitionwould slow but not stop the development of molecular genetic techniques and the biotech-no l ogy i ndus t r y because th ere are severa] good

alternatives for maintaining exclusive control of biological inventions. Development would be

slowed primarily because information thatmight otherwise become public would bewithheld as trad e secrets. A m ajor consequencewould be that desirable products would takelonger to reach the market.

Alternatively, Congress could overrule theSup reme Court’s decision by am ending th e pat-ent law to p rohibit patents on organisms otherthan the plants covered by the two statutesmentioned in option C. This would d emonstratecongressional intent that living organisms couldbe patented only by specific statute.

E. Congress could pass a comprehensive law cov-ering any or all organisms (except humans).

This option recognizes that Congress candraw lines where it sees fit in this area. It couldspecifically limit patenting to micro-organisms,




or it could encourage the breeding of agricul-turally important animals by granting patentrights to breeders of new and distinct breeds. Inthe interest of comprehensiveness and uniform-ity, one statute could cover plants and all otherorganisms that Congress desires to be patent-able.

Genet ics and soc ie ty

FINDINGS

Continued advances in science and technol-ogy are beginning to provide choices that strainhuman value systems in areas where previouslyno choice was possible. Existing ethical andmoral systems do not provide clear guidelinesand directions for those choices. New program s,both in public institutions and in the popularmedia, have been established to explore the

relationships among science, technology, socie-ty, and value systems, but more w ork needs tobe done.

Genetics-and other areas of the biologicalsciences—have in comm on a mu ch closer rela-tionship to certain ethical questions than domost advances in the physical sciences orengineering. The increasing control over the

characteristics of organisms and the potentialfor altering inheritance in a directed fashionraise again questions about the relationship of hu man s to each other and to other living things.People respond in different ways to this poten-tial; some see it (like many predecessor develop-ments in science) as a challenging opportunity,others as a threat, and still others respond withvague u nease. Although man y people cannot ar-ticulate fully the basis for their concern, ethical,moral, and religious reasons are often cited.

The public’s increasing concern about the ad-vance of science and impacts of technology hasled to dem and s for greater pa rticipation in deci-sions concerned with scientific and technologi-cal issues, not only in the United States butthroughout the world. The demands imply newchallenges to systems of representative govern-ment. In every Western country, new mecha-

nisms h ave been devised for increasing citizenparticipation.

The public has already become involved indecisionmaking with regard to genetics. As thescience develops, ad ditional issues in wh ich thepu blic will demand involvement can be antici-pated for the years ahead. The question then be-comes one of how best to involve the p ublic indecisionmaking.

I s s u e s a n d O p t i o n s — G en e t i c s an d So c i e ty

Issue: H ow shou ld the pub l i c be in -volved in determining policy re-lated to new applications of ge-netics?

Because public demands for involvement areunlikely to diminish, ways to accommodatethese demand s must be considered.

OPTIONS:

A.

B.

Congress could specify that public opinion must be sought in formulating all major pol-icies concerning new applications of genetics,including decisions on the funding of specific

research projects. A “Public ParticipationStatement” could be mandated for all such

decisions.

Congress could maintain the status quo, allow-

ing the public to participate only when it decides to do so on its own initiative.

If option A were followed , there would be nocause for claiming that public involvement wasinadequate (as occurred after the first set of Guidelines for Recombinant DNA Research wasprom ulgated). Option A p oses certain problems:How to identify a major policy and at what stagepu blic involvement wou ld be required. Shou ldit take place only wh en technological develop-ment and application are imminent, or at thebasic research stage?

Option B would be less cumbersome to effect.It would permit the establishment of ad hocmechanisms when necessary.



—

Ch. 1—Summary: Issues adoptions . 25

ISSUE:

There

too little

H o w c a n t h e l e v e l o f p u b l i cknowledge concern ing gene t icsand its potential be raised?

are some educators who believe thattime is spent on genetics within the

traditional educational

s y s t e m . o u t s i d e t h etrad itional school system, a nu mber of sourcesmay contribute to increased public understand-ing of science and the relationship betweenscience and society.

Efforts to increase public understandingshould , of course, be combined with carefullydesigned evaluation p rograms so that the effec-tiveness of a pr ogram can be assessed.

OPTIONS:

A. Programs could be d e v e l o p e d t o i n c r e a s e

public understanding of science and the relationship between science, technology, and

society.

Public understanding of science in today's

world is essential, and there is concern about

the adequacy of the public 's knwledge.

B. programs could be established to monitor the

level of public understanding of genetics and of science in general, and to determine wheth-

er public concern with decisionmaking in

science and technology is increasing.

Selecting this option would indicate that

there is need for additional information, andthat Congress is interested in involving thepublic indeveloping science policy.

C. The copyright laws could be amended to per-

mit schools to videotape television programs

for educational purposes.

Und er current copyright law, videotaping tel-evision programs as they are being broadcastmay infringe on the rights of the program’sown er, generally its prod ucer. The legal statusof such tap es is presently the su bject of litiga-tion.

In favor of this option, it should be noted th atmany of the programs are made at least in partwith p ublic fund s. Removing the copyright con-straint on schools would make these programsmore available for another public good, educa-

tion. On the other hand , this option could h avesignificant economic consequences to the copy-right owner, whose market is often limited toeducational institutions.

ISSUE: Should Congress begin prepar -

ing now to reso lve i ssues tha thave not yet aroused much pub-lic debate but which may in thefu ture?

As scientific understanding of genetics andthe ability to manipulate inherited character-istics develops, society may face some d ifficultquestions that could involve trad eoffs betweenindividual freedom and the needs of society.This will be increasingly the case as genetictechnologies are applied to humans. Develop-ments ar e occurring rap idly. Recombinant DNA

technology was developed in the 1970’s. In thespring of 1980, investigators succeeded in thefirst gene rep lacement in mam mals; in th e fallof 1980, the first gene substitution in humanswas attempted.

Although this study was restricted to nonhu-man applications, many people assume fromthese and other examples that what can be donewith lower animals can be done w ith hum ansand will be. Therefore, some action might betaken to better prep are society for decisions onthe application of genetic technologies to

humans.

OPTIONS:

A.

B.

A commission could be established to identify

central issues, the probable time frame for ap-

plication of various genetic technologies to

humans, and the probable effects on society,

and to suggest courses of action. The commis-

sion might also consider the related area of

how participatory democracy might be combined with representative democracy in deci-

sionmaking.

The life of the President Commission could

be extended for the study of Ethical problems

in Medicine and Biomedical and Behavioral Re-

search, for the purpose of addressing these

issues.



26 . Impacts of Applied Genetics—Micro-Organisms, plants, and Animals

This 1 l-mem ber ‘Comm ission w as establishedin November 1978 and terminates on December31, 1982. It could be asked to broaden its cover-age to additional areas. This would require thatthe life span of the commission be extended andadd itional funds be appropriated.

A potential disadvan tage to using the existingcommission to address societal issues associatedwith genetic engineering is that a number of

issues already exist, and more are likely to arisein the years ahead. Yet there are also otherissues in medicine and biomedical and behav-ioral research n ot associated with genetic engi-neering that also need review. Whether allthese issues can be addressed by one commis-

sion should be considered. Comments from theexisting comm ission w ould assist in d eciding th emost ap prop riate course of action.



C h a p t e r 2

I n t r o d u c t i o n



C h a p t e r 2

Page

The Origins of Genetics. . . . . . . . . . . . . . . . . , . . 29

(Genetics in the 20th Century. . . . . . . . . . . . . . . . 33

The Ridd le of the Gene , . . . . . . . , . . . . . , . . . . . 33The Genetic Cod e . . . . . . . . . . . . . . . . . . . . . . . , .37Developing Genetic “technologies . . . . . . . , . . . . 39The Basic Issues . . . . . . . . . . . . . . . . . . . . . . . . . 43

How Will Applied Genetics Be Used? . . . . . . . . 43What Are th e Dangers? . . . . . . . . . . . . . . . , . . 43

Figures

Figure No. Page

5. The Inheritance Pattern of Pea Color. . . . . , , 306. Chromosom es . . . . . , . . . . . . . . . . . . . . . . . . 32

7. The Gr iffith Experiment . . . . . . . . . . . . . . . . 348. The Structure of DNA . . . . . . . . . . . . . . . . . 369. Replicat ion of DN A . . . . . . . . . . . . . . , . . . . 37

10. The Genetic Code . . . . . . . . . . . . . . . . . . . , 3811. The Expression of Genetic Information in

the Cell . . . . . . . . . . . . . . . . . . . . . . . . . . , 3912. Transduction: The Transfer of Genetic

Material in Bacteria by Means of Viruses . . . . 39

13. Conjugation: The Transfer of GeneticMa teri al in Bacteria b y M atin g. . . . . . . . . . . . 40

14. Recombinant DNA: The Technique of

Recombining Genre From One Species WithThose From Another , ! . . . . . . . , . . . . . . . . . 41

15. An Example of How the Recombinant DNA

Technique May Be Used To Insert NewGenes Into Bacterial Cells. . . . . . . . . , . . . . . 42



Ch a p t e r 2


Hu mankind is gaining an increasing un der-standing of heredity and variation am ong livingthings—the science of genetics. This report ex-amines both the critical issues arising from thescience and technologies that spring from ge-netics, and the potential impacts of these ad-vances on society. They are the most rapidlyprogressing areas of human knowledge in theworld today.

Genetic technologies exist only within thelarger context of a maturing science. The key toplanning for their potential is understandingnot simply a particular technology, or breedingprogram, or new opportunity for investment,but h ow the field of genetics works and how it

interacts with society as a whole.

The technologies that this report assesses canbe expected to have pervasive effects on life inthe future. They touch on th e most fundam en-tal and int imate needs of mankind : health care,supplies of food and energy, and reproduction.At the same time, they trigger concerns in areas

T h e o r i g i n s o f g e n e t i c s

For the past 10,000 years, a period encom-passing less than one-half of 1 percent of man’s

time on Earth, the human race has developed

under the impetus of applied genetics. As tech-niques for planning, cult ivating, and storing

crops replaced subsistence hunting and forag-

ing, the character of humanity changed as well.

From the domestication of animals to the devel-

opment of permanent settlements, from the rise

o f m odern sc i ence to the daw n o f b io t ech-

nology, the genetic changes that mankind has

directed have, in turn, affected the nature of his

society.

Applied genetics depends on a fundamental

principle–that organisms both resemble and

differ from their parents. It must have required

great faith on the part of Neolithic man to bury

equally as important: the dwindling supplies of natural resources, the risks involved in basicand applied scientific research and develop-ment, and the natu re of innovation itself.

As always, some decisions concerning the useof the new technologies will be made by themarketp lace, wh ile others will be made by var-ious institutions, both pu blic and private. In thecoming years, the p ublic and its repr esentativesin Congress and other governm ental bodies willbe called on to make difficult decisions becauseof society’s knowledge about genetics and itscapabilities.

This report d oes not make recommendationsnor does it attemp t to resolve conflicts. Rather,it clarifies the bases for making judgments bydefining the likely impacts of a group of technol-ogies and tracing their economic, societal, legal,and ethical imp lications. The new genetics w illbe influential for a long time to come. Althoughit will continu e to change, it is not too early tobegin to monitor its course.

perfectly good grain during one season in thehope of growing a new crop several monthslater–faith not only that the seed w ould ind eedreturn, but tha t it would d o so in the form of thesame grain-producing crop from which it hadsprung. This permanence of form from onegeneration to the next has been scientificallyunderstood only within the past century, butthe understanding has transformed vague be-liefs in the inheritance of traits into the scienceof genetics, and rule-of-thum b animal an d plantbreeding into the modern manipulations of genetic engineering.

The major conceptual boost for the scienceof genetics required a shift in perspective,from the simple observation that characteristics

29



30 . lmpacts of Applied Genetics—Micro-Organisms, Plants, and Animals

passed from parents to offspring, to a study of the un derlying agent by w hich this transmissionis accomplished. That shift began in the gardenof Gregor Mendel, an obscure monk in mid-19thcentury Austria. By analyzing generations of controlled crosses between sweet pea plants,

Mendel was able to identify the rudimentarycharacteristics of what was later termed thegene.

Mendel reasoned that genes were the vehicleand repository of the hereditary mechanism,and that each inherited trait or function of anorganism had a specific gene directing its devel-opment and appearance. An organism’s observ-able characteristics, functions, and measu rableproperties taken together had to be based some-how on the total assemblage of its genes.

Mendel’s analysis showed that the genes of

his pea plants remained constant from one gen-eration to the next, but more importantly, hefound that genes and observable traits were notsimply matched one-for-one. There were, infact, two genes involved in each trait, with asingle gene contributed by each parent. Whenthe genes controlling a p articular trait are iden-tical, the organism is homozygous for that trait;if they are not, it is heterozygous.

In the Mend elian crosses, homozygou s plantsalways retained the expected characteristics.But heterozygous plants did not simply display a

mixture of their d ifferent genes; one of the tw otended to p redominate. Thus, when homozy -gous yellow-seed peas w ere crossed w ith homo-zygous green-seed p lants, all the offspring w erenow heterozygous for seed color, possessing a“green” gene from one p arent and a “yellow”from the other. Yet all of them tu rned out to beindistingu ishable from the yellow-seed par ent:Yellow-seed color in peas was dominant togreen.

But even though the offspring resembledtheir dominant p arent, they could be show n tocontain a genetic difference. For w hen the het-

erozygotes were now crossed w ith each other, acertain number of recessive green-seed plantagain appeared among the offspring. This oc-curred whenever an offspring was endowedwith a p air of genes that was homozygous for

the green-seed trait—and it occurred at a rateconsistent with the random selection of one of two genes from each parent for passage to thenew generation. (See figure 5.)

Genes were real—Mendel’s work made thatclear. But where were they located, and what

were they? The answer, lay within the nucleusof the cell. Unfortunately, most of the contentsof the nucleus were u nobtainable by biologistsin Mendel’s time, so his published findings wereignored. Only during the last decades of the19th century did improved microscopes andnew dyes permit cells to be observed with anacuity never before possible. And only by the

Figure 5.-The Inheritance Pattern of Pea Color

Y = yellow gene g = green gene

Each parent contributes only one seed-color gene to the off-spring. When the two YY and gg homozygotes are crossed,the genetic composition of all offspring is Yg:

OffspringAll Yg offspring are heterozygous, and all have yellowseeds, indicating that the Y yellow gene is dominant overthe g green gene.

When these Yg heterozygotes are crossed with each other:

Thus, 3/4 of these offspring will have yellow seeds, but theirindividual genetic composition, YY of Yg, maybe different.




Ch. 2—Introduction q 37

beginning of the 20th century did scientists rediscover Mendel’s work and begin to appre-

ciate fully the significance of the cell nucleus

and its contents.

Even in the earliest microscopic studies,however, certain cellular components stoodout; they were deeply stained by added dye. Asa result, they w ere du bbed ‘(colored bod ies, ” orchromosomes. Chromosomes were seen rela-tively rarely in cells, with most cells showing just a central dark nucleus surrounded by anextensive light grainy cytoplasm. But periodi-cally the nu cleus seemed to d isappear, leavingin its place long thready material that con-solidated to form the chromosomal bodies. (Seefigure 6a.) Once formed, the chromosomesassembled along the m idd le of the cell, copiedthemselves, and then moved ap art wh ile the cell

pinched itself in half, trapping one set of chromosomes in each of the two halves. Thenthe chromosomes themselves seemed to dis-solve as two new nuclei appeared, one in eachof the tw o new ly formed cells. (See figure 6b. )

Thus, the same number of chromosomes ap-peared in p recisely the same form in every cellof an organism except the germ, or sex, cells.Furthermore, the chromosomes not only re-mained constant in form and num ber from onegeneration to the next, but were inherited inpairs. They were, in short, manifesting all the

traits that Mendel had prescribed for genesalmost th ree decades earlier. By the beginningof the 20th century, it was clear that chromo-somes were of central importance to the life his-tory of the cell, acting in some unspecified man-ner as the vehicle for the Mend elian gen e.

If this conclusion was strongly implied by theevents of cell d ivision, it became obvious wh enreproduction in whole organisms was analyzed.It had been established by the latter part of the19th century that the germ cells of plants andanimals—pollen and ovum , sperm and egg—ac-tually fuse in the process of fertilization. Germcells d iffer from other bod y cells in on e impor-tant respect—they contain only half the usualnumber of chromosomes. This chromosomehalving within the cell was apparently donevery precisely, for every sperm and egg con-

tained exactly one representative from eachchromosome pair. When the two germ cellsthen fused during fertilization, the offspringwere supplied with a fully reconstituted chro-mosome complement, half from each parent.Clearly, chromosomes were the material link from one generation to the next. Somewherelocked within them was the substance of bothheredity—the fidelity of traits betw een genera-tions; and diversity—the potential for geneticvariation and change.




Figure 6.— Chromosomes

6a. An example of

Step 1

Photo credit: Professor Judith Lengyel, Molecular Biology institute, UCLA

Optical micrograph of chromosomal material from the salivary gland of the larva of the

common fruit fly, Drosophila melanogaster

a chromosome body from a higher organism.

Step 3 Step 4

6b. In Step 1, the chromosome bodies are still uncondensed.

Step 5 Step 6

In Steps 2 and 3, the chromosomes condense into thread-like bodies and align themselves near the center of the cell.In Steps 4 and 5, the chromosomes begin to separate and are pulled to the opposite poles of the cell.

In Step 6, the chromosomes return to an uncondensed state and the cell begins to constrict about the middle to formtwo new cells.




Ch. 2—introduction . 33

( k i n e t i c s i n t h e 2 0 t h c e n t u r y

During the first few d ecades of the 20th cen-tury, scientists searched for progressivelysimpler experimental organ isms to clarify p ro-

gressively more complex genetic concepts. Firstwas Thomas Hunt Morgan’s Drosophila—gnat-sized fruit flies with bulbous eyes. These insectshave a simple array of four easily distinguish-able chromosome pairs per cell. They repro-duce rapidly and in large numbers under thesimplest of laboratory conditions, supplying anew generation every month or so. Thus, re-searchers could carry out an enormous numberof crosses employing a whole catalog of dif-ferent fruit fly traits in a relatively brief time.

It became obvious from the extensive Dros-

ophila data that certain traits were more likelyto be inherited together than others. Yellowbodies and ruby eyes, for instance, almost al-ways w ent together, with both in turn , appear-ing more frequently than expected with thetrait known as “forked bristles. ” All three traits,however, showed up only randomly withcurved w ings. Certain genes thus seemed to belinked to one another. The entire Drosophilagenome, in fact, fell into four distinct linkagegroups. The physical basis for these groups, notsurprisingly, consisted of the four fruit flychromosomes. Linked genes behaved as they

did because they were located on the samechromosome.

Soon, scientists learned that they could notonly assign particular genes to particular Droso-

phila chromosomes bu t could identify the rela-tive locations of different genes on a givenchromosome. This gene mapping was possible

T h e r i d d l e o f t h e g e n e _

because linkage itself was not permanent,linked genes sometimes separ ated. For instance,wh ile yellow bodies, ruby eyes, and forked bris-

tles were all linked traits, the first two stayedtogether far more frequently than either didwith the third.

The degree of linkage between two genes w ashypothesized to be directly proportional to thedistance between them on the chromosome,mainly because of a unique event that occursduring the development of germ cells. Beforethe normal chromosome num ber is halved, thechromosomes crowd together in the center of the cell, coiling tightly around each other, prac-tically fusing along their entire length. It is in

this state that crossing-over (or natural recombi-nation)—the actual p hysical exchange of partsbetween chromosomes—occurs. No chromo-some emerges from the exchange in the samecondition as before; the lengths of chromo-somes are reshuffled before being transferredto the n ext generation.

The idea of linkage m eant that Mend el’s for-mulations had to be modified. Clearly, geneswere not completely independent units. Furtherwork with Drosophila in the 1920’s showed thatgenes were also not permanent and couldchange over time. Although natu ral mutations

occurred at a very slow rate, exposing fruit fliesto X-rays accelerated their frequency enor-mously. Exposure of a parental fly populationled to an array of new traits among their off-spring—traits which, if they were neither lethalnor sterilizing, could be passed from one gen-eration to the next.

With all this research, nobody yet knew wh attriguing observations m ade a d ecade earlier by

the gene was made of. The first evidence that a British physician, Fred Griffith. He hadit consisted of deoxyribonucleic acid (DNA) worked with tw o types of pneum ococcus (the

emerged from the work of Oswald Avery, Colin bacteria responsible for pneumonia) and withMacLeod, and Maclyn McCarty at the Rockefel- two different bacteria within each type. Oneler Institute in New York in the early 1940’s. bacterium in each type was coated in a polysac-Avery’s group took as its starting p oint some in- charide capsule; the other was bare. Bare bac-




teria gave rise only to bare p rogeny, wh ile thosewith capsules produced only encapsulatedforms. Only the encapsulated forms of bothtypes II and III could cause disease; bare bac-teria were benign. (See figure 7a.) But whenGriffith took some encapsulated type III bacteria

that had been killed and rendered harm less andmixed them with bare bacteria of type II, thepresumably safe mixture became virulent: Miceinjected w ith it died of a massive pneum onia in-fection, Bacteria recovered from these animalswere found to be of type 11—the only living bac-teria the mice had received—now wrapped intype III capsules. (See figure 7b.)

Avery’s group recognized Griffith’s find ing asa genetic phenom enon; the dead type 111 bacte-ria must have delivered the gene for makingcapsules into the genetic complement of the

living typ e 11 recipients. By meticulous research ,Avery’s group found that the substance whichcaused the genetic transformation w as DNA.

It had been in 1868, just 3 years after Mendelhad published his findings, that DNA was dis-covered by Friedrich Miescher. It is an extreme-ly simple molecule composed of a small sugarmolecule, a phosphate group (a phosphorousatom surroun ded by four oxygen atoms), andfour kind s of simple organic chemicals know nas nitrogenous (nitrogen-containing) bases. To-gether, one sugar, one phosphate, and one baseform a nucleotide—the basic structural unit of

the large DNA molecule. Because it is so simple,DNA had appeared to be little more than amonotonou s conglomeration of simp le nucleo-tides to scientists in the early 20th century. Itseemed unlikely that such a prosaic moleculecould direct the appearance of genetic traitswhile faithfully reproducing itself so that in-formation could be transferred between gen-erations. Although Avery’s results seemed clearenough, many were reluctant to accept them.

Those doubts were finally laid to rest in abrief report published in 1953 by James Watson

and Francis Crick. By using X-ray crystallo-graphic techniques and building complex mod-els—and withou t ever having actually seen themolecule itself—Watson and Crick reported thatthey had discovered a consistent scientificallysound structure for DNA.

Figure 7.—The Griffith Experiment

7a. There are two types of pneumococcus, each of whichcan exist in two forms:

where R represents the rough, nonencapsulated, benignform; and

S represents the smooth, encapsulated, virulentform.

7b. The experiment consists of four steps:

Virulent strain (1)

Mice injected with nonvirulent Rll do not become infected.

Virulentstrain,

heat-killed

(3)

The virulent SIII is heat-killed. Mice injected with it do notdie.

heat-killed II

(4)When mice are injected with the nonvirulent RII and theheat-killed SIII, they die. Type II bacteria wrapped in type Illcapsules are recovered from these mice.





The structure that Crick and Watson uncov-ered solved part of the genetic pu zzle. Accord-ing to them, the phosphates and sugars formedtwo long chains, or backbones, with one nitrog-enous base attached to each sugar. The twobackbones were held together like the supports

of a ladder by weak attractions between thebases protruding from the sugar molecules. Of the four d ifferent nitrogenou s bases—aden ine,thym ine, guanine, and cytosine—attractions ex-isted only between adenine(A) and thymine(T),and between guan ine(G) and cytosine(C). (Seefigure 8a) Thus, if a stretch of nucleotides onone backbone ran:

the other backbone had to contain the directlyopposite complementary sequence:

T-A-G-A-A-T-T. . .

The complementary pairing between bases run-ning d own the center of the long molecule wasresponsible for holding together the two other-wise independent chains. (See figure 8b. ) Thus,the DNA molecule was rather like a zipper, withthe bases as the teeth and the sugar-phosphatechains as the strands of cloth to which each zip-per h alf was sew n. Crick and Watson also foundthat in the presence of water, the two poly -nucleotide chains did not stretch out to fulllength, but twisted arou nd each other, formingwhat has un dou btedly become the most glori-

fied structure in the history of biology-the dou-ble helix. (See figure 8c.)

The stru cture w as scientifically elegant. But itwas received enthusiastically also because it im-plied how DNA w orked. As Crick and Watsonthemselves noted:

If the actual order of the bases on one of thepair of chains were given, one could write downthe exact order of the bases on th e other one,because of th e specific pairing. Thus one chainis, as it were, the complement of the other, andit is this feature which suggests how the desoxy-ribonucleic acid molecule might d up licateitself. ’

When a double-stranded DNA molecule is un-zipped, it consists of two separate nucleotidechains, each with a long stretch of unpairedbases. In the presence of a mixture of nucleo-tides, each base attracts its complementarymatch in accordance with the inherent affinitiesof adenine for thymine, thymine for adenine,guan ine for cytosine, and cytosine for gu anine.The result of this replication is tw o DN A m ole-cules, both precisely identical to each other andto the original molecule—which explains the

faithful duplication of the gene for passage fromone gen eration to t he next. (See figure 9.)

Crick and Watson’s work solved a major rid-dle in genet ic research. Because George Beadleand Edward Tat urn had recently discoveredthat genes control the appearance of specificproteins, and that one gene is responsible forprod ucing one specific protein, scientists nowknew w hat the genetic material was, how it rep-licated, and wh at it produ ced. But they had yetto determine how genes expressed themselvesand produ ced proteins.

‘James D. Watson and Francis Crick, %enetical Implications of

the Structures of Deoxyribose Nucleic Acid, ” Nature 171, 1953. pp.

737-8.




A

Adenine

Figure 8.—The Structure of DNA

o-o”

5

0 .-o”

8a. The pairing of the four nitrogenous bases of DNA:

Adenine (A) pairs with Thymidine (T)Guanine (G) pairs with Cytosine (C) -o”

o .

8b. The four bases form the four letters in the alphabet ofthe genetic code. The sequence of the bases along thesugar-phosphate backbone encodes the genetic in-formation.

A schematic diagram of the DNA double helix. A three-dimensional representation of the DNA double helix.

&. The DNA molecule is a double helix composed of two chains. The sugar-phosphate backbones twist around the out-

side, with the paired bases on the inside serving to hold the chains together.





T h e g e n e t i c

Figure 9.—Replication of DNA

Old Old

Old New New Old

When DNA replicates, the original strands unwind and

serve as templates for the building of new complementary

strands. The daughter molecules are exact copies of theparent, with each having one of the parent strands.


c o d e

Proteins are the basic materials of cells. Some Ironically, proteins are far more complex andproteins are enzymes, wh ich catalyze reactions diverse than the four nucleotides that helpwithin a cell. In general, for every chemical re- create them. Proteins, too, are long chains madeaction in a living organism, a specific enzyme is up of small units strung together. In this case,required to trigger the process. Other proteins how ever, the un its are amino acids rather thanare structural, comprising most of the raw ma- nucleotides—and there are 20 different kind s of terial that forms cells. amino acids. Since an average protein is a few

76-565 0 - 81 - 4




hundred amino acids in length, and since anyone of 20 amino acids can fill each slot, the num -her of p ossible proteins is enormous. N everthe-less, each protein requires the strictest orderingof amino acids in its structure. Changing asingle amino acid in the entire sequence can

drastically change the protein’s character.It was now possible for scientists to move

nearer to an appreciation of how genes func-tioned. First had come the recognition that DNAdetermined protein; now it was evident that thesequence of nucleotides in DNA determined alinear sequence of amino acids in proteins.

By the early 1980’s, the way proteins were

manufactured, how their synthesis was regu-lated, and the role of DNA in both processeswere understood in considerable detail. Theprocess of transcribing DNA’s message-carry-ing the message to th e cell’s miniatu re p roteinfactories and building proteins-took place

through a complex set of reactions. Each aminoacid in the protein chain was represented bythree nucleotides from the DNA. That three-base unit acted as a word in a DNA sentencethat sp elled ou t each protein-the genetic code.(See figure 10.)

Through the genetic code, an ent ire gene—alinear assemblage of nu cleotides-could now be

THIRD BASE

Figure 10.—The Genetic Code

THIRD BASE

SECOND THIRD BASE THIRD BASE

q och (ochre); amb (ambert, and end are step signals for translation, i.e.,signals the end of synthesis of the protein chain.

Amino acid alaninearginineasparagineaspartic acidasn and/or aspcysteineglutamineglutamic acidgin and/or gluglycinehistidineisoleucineIeucineIysinemethioninephenylaianine

prolines e r i n ethreoninetryptophantyrosinevaline

symbol alaargasnaspasxCysglngluglxglyhisileuIeuIysmetphe

proserthrtrptyrval

Each amino acid is determined by athree letter code (A, G, T, or C) alongthe DNA. If the first letter in the codeis A, the second is T, and third is A,the amino acid will be tyrosine (or tyr)in the complete protein molecule. ForIeucine (or Ieu), the code is GAT, and

so forth. The dictionary above givesthe entire code,




Ch. 2—introduction q 39

read like a book. By the 1970’s, researchers had Figure 11.—The Expression of Genetic Information

learned to read the code of certain proteins, in the Cell

synthesize their DNA, and insert the DNA intobacteria so that the protein could be p roduced.(See figure 11.)

Meanwhile, other scientists were studying

the genetics of viruses and bacteria. The com-bination of these stud ies with those investigat-ing the genetic code led to the innovations of genetic engineering.

Deve lop ing

In the early 1960’s,


g e n e t i c t e c h n o l o g i e s

scientists discovered ex- DNA along with it . Thus, when the new virusactly how genes move from one bacterium toanother. One such mechanism uses bacterio-

phages-viruses that infect bacteria-as inter-mediaries. Phages act like hypodermic needles,injecting their DNA into bacterial hosts, whereit resides before being passed along from one

generation to the next as part of the bacterium’sown DNA. Sometimes, however, the injectedDNA enters an active ph ase and p rodu ces a cropof new virus particles that can then burst out of their host. Often during this process, the viralDNA inadvertently takes a piece of the bacterial

particles now infect other bacteria, they bringalong several genes from their previous host.

This viral transduction-the transfer of genesby an intermediate viral vector or vehicle-could be used to confer new genetic traits onrecipient bacteria. (See figure 12. )

Bacteria also transfer genes directly in a proc-ess called ’conjugation, in which one bacteriumattaches small projections to the surface of anearby bacterium. DNA from the donor bacte-rium is then passed to the recipient through the

Figure 12.—Transduction: The Transfer of Genetic Material in Bacteria by Means of Viruses

Bacterium

BacterialEmpty chromosome

viral coat fragments. New virus

bacterium

In step 1 of viral transduction, the infecting virus injects its DNA into the cell. In step 2 when the new viral particles areformed, some of the bacterial chromosomal fragments, such as gene A, may be accidently incorporated into these progenyviruses instead of the viral DNA. In step 3 when these particles infect a new cell, the genetic elements incorporated from thefirst bacterium can recombine with homologous segments in the second, thus exchanging gene A for gene a.

SOURCE: Office of Technology Assessment




projections. The ability to form projections anddon ate genes to neighbors is a genetically con-trolled trait. The genes controlling this trait,how ever, are not located on the bacterial chro-mosomes. Instead, they are located on separategenetic elements called plasmids-relatively

small molecules of double-stranded DNA, ar-ranged as closed circles and existing autono-mously within the bacterial cytoplasm. (Seefigure 13.)

Plasmids and phages are two vehicles—orvectors-for carrying genes into bacteria. Assuch, they became tools of genetic engineering;for if a specifically selected DNA could be intro-du ced into these vectors, it would then be pos-sible to tran sfer into bacteria the bluep rints forproteins—the building blocks of genetic charac-teristics.

But bacteria had been confronting the inva-sion of foreign DN A for millennia, and they hadevolved protective mechanisms that preservedtheir own DNA while destroying the DNA thatdid not belong. Bacteria survive by producingrestriction enzymes. These cut DNA moleculesin places where specific sequences of nucleo-tides occur—snipping the foreign DNA, yet leav-ing the bacteria’s own genetic complementalone. The first restriction enzyme that was iso-

lated, for instance, would cut DNA on ly when itlocated the sequ ence:

(G-A-A-)

( C - T - T )

If the sequence occurred once in a circular plas-

mid, the effect would simply be to open thecircle. If the sequence were repeated severaltimes along a length of DNA, the DNA w ould bechopped into several small pieces.

By the late 1970’s, scores of different restric-tion enzymes had been isolated from a varietyof bacteria, with each enzym e having a u niquespecificity for one specific nucleotide sequence.These enzymes were another key to genetic en-gineering: they not only allowed plasmids to beopened u p so that new DN A could be inserted,but offered a way of obtaining manageablepieces of new DNA as well. (See figure 14.)

Using restriction enzymes, almost any DNAmolecule could be snipped, shaped, andtrimmed with pr ecision.

Cloning DNA—that is obtaining a large quanti-ty of exact copies of any chosen DN A m oleculeby inserting it into a host bacterium-becametechnically almost simple. The piece in questionwas m erely snipp ed from th e original molecule,inserted into the vector DNA, and p rovided w ith

Figure 13.—Conjugation: The Transfer of Genetic Material in Bacteria by Mating

Plasmid

In conjugation, a plasmid inhabiting a bacterium can transfer the bacterial chromosome to a second cell where homologoussegments of DNA can recombine, thus exchanging gene B from the first bacterium for gene b from the second.




Ch. 2—introduction q 4 1

Figure 14.—Recombinant DNA: The Technique ofRecombining Genes From One Species

With Those From Another

New DNA

amount of DNA protein

Restriction enzymes recognize certain sites along the DNA

and can chemically cut the DNA at those sites. This makes

it possible to remove selected genes from donor DNA mole-

cules and insert them into plasmid DNA molecules to form

the recombinant DNA. This recombinant DNA can then becloned in its bacterial host and large amounts of a desired

protein can be produced.


a bacterial host as a suitable environment forreplication. The d esired piece of DNA could berecombined with a plasmid vector, a procedure

that gave r ise to recombinant DN A (rDNA),aIso

known as gene splicing. Since bacteria can begrown in vast quantities, this process couldresult in large-scale production of otherwisescarce and expensive proteins.

Although placing genes inside of bacteria isnow a relatively straightforward procedure, ob-taining precisely the right gene can be difficult.Three techniques are currently available:

q Ribonucleic acid—RNA—is the vehiclethrough which the message of DNA is readand transcribed to form proteins. The RNA

that carries the message for the desiredprotein is first isolated . An enzym e, called‘reverse transcriptase, is then ad ded to theRNA. The enzyme triggers the formation of DNA—reversing th e norm al process of pro-tein prod uction. The DN A is then inserted

q

q

into an appropriate vector. This was theprocedu re used to obtain th e gene for hu -man insulin in 1979. (See figure 15. )The gene can also be synthesized, orcreated, directly, since the nucleotide se-quence of the gene can be deduced from

the amino acid sequence of its proteinprod uct. This procedu re has w orked w ellfor small proteins—like the growth regu-latory hormone somatostatin—which haverelatively short stretches of DNA coding.But somatostatin is a tiny protein, only 14amino acids long. With three nucleotidescoding for each amino acid, scientists hadto synthesize a DNA chain 42 nucleotideslong to prod uce the comp lete hormone. Forlarger proteins, the gene-synthesis ap-proach rapidly becomes highly impractical.The third method is also the most con-

troversial. In this “shotgun” app roach, theentire genetic complement of a cell ischopped up by restriction enzymes. Eachof the DNA fragments is attached next tovectors and transferred into a bacterium;the bacteria are then screened to find th osemaking the desired product. Screeningthou sand s of bacterial cultu res was part of the technique tha t enabled the isolation of the hu man interferon gene. *

At present, these techniques of recombina-tion w ork mainly with simple m icro-organisms.

Scientists have only recently learned how to in-troduce novel genetic material into cells of higher plants and animals. These higher cellsare being ‘engineered’ in totally different ways,by grow ing p lant or animal cells in ‘tissue cul-ture’ systems, in vitro.

Tissue culture systems work with isolatedcells, with entire pieces of tissue, and to a farmore limited extent, with whole organs or even

early embryos. The techniques make it possibleto manipulate cells experimentally and undercontrolled conditions. Several techniques areavailable. For example, in one set of experi-

ments, complete plants have been grown fromsingle cells—a breakthrough that may permit

“Strictly speaking, RNA was transcribed using the shotgun

approach into DNA, which was then cloned into bacteria andscreened.




Figure 15.-An Example of How the Recombinant DNA Technique May Be UsedTo Insert New Genes Into Bacterial Cells

The first part of the technique involves the manipula- 11. A bacterial plasmid, which is a small piece of circular

tions necessary to isolate and reconstruct the desired DNA, serves as the vehicle for introducing the new gene

gene from the donor: (obtained in part I above) into the bacterium:‘a)

b)

c)

The RNA that carries the message (mRNA) for the. . ,

a) The circular plasmid is cleaved by the appropriatedesired protein product is isolated. restriction enzyme.

The double-stranded DNA is reconstructed from the b) The enzyme terminal transerase extends the DNAmRNA. strands of the broken circle with identical basesin the final step of this sequence, the enzyme ter- (four cytosines in this case, to allow complemen- minal transferase acts to extend the ends of the

DNA strands with short sequences of identical

bases (in this case four guanines).

IMessenger RNA

from animal cell

a)

Enzymatic

reconstruction

Double-strand DNA t

b)

I

Terminaltransferase

Ill. The final product, a bacterial plasmid containing the

new gene, is obtained. This plasmid can then be in-

serted into a bacterium where it can be replicated andproduce the desired protein product:

a)

b)

tary base pairing with the guanines added to the

gene obtained in part i).

The gene obtained in part i and the plasmid DNAfrom part ii are mixed together and anneai

because of the complementary base-pairing be-tween them.Bacterial enzymes fill in any gaps in the circle,

sealing the connection between the plasmid DNA

and the inserted DNA to generate an intact cir-

cular plasmid now containing a new gene.

Uptake by cell;

Ill repair by

II

Bacterial plasmid DNA

a)

enzymes




Ch. 2—introduction q 43

hu nd reds of plants to be grow n asexually froma small sample of plant material. Just as withbacteria, the cells can be induced to take uppieces of DNA in a process call transformation.They can also be exposed to mutation-causingagents so that they produce mutants with de-

sired pr operties. in another set of experimen ts,two different cells have been fused to form anew, single-cell “hybrid” that contains thegenetic complements of both antecedents. Inboth cases, the success of tissue culture and cell

T h e b a s i c i s su e s

App lied genetics is like no otherBy itself, it may enable tremend ous

fusion* can be used to direct efficient, fast ge-netic changes in plants. (See ch. 8.)

Cell culture techniques, while not strictly ge-netic manipulation, form a m ajor aspect of mod -ern biotechnology. Combined with genetic ap-proaches, their potential is only on the verge of

being realized.

q A related technique is protoplasm fusion, or the fusion of cells

whose walls have been removed to leave only membrane-boundcells. The cells of bacteria, fungi, and plants must all be freed of

their walls before they can be fused.

technology.adv ances in

conquering diseases, increasing food produ c-

tion, prod ucing new and cheaper indu strial sub-stances, cleaning up pollution, and understand-ing the fundamental processes of life. Becausethe technology is so powerful, and because it in-volves the basic roots of life itself, it carries withit potential hazard s, some of which might arisefrom basic research, others of wh ich m ay stemfrom its applications.

As the impacts of genetic technologies are dis-cussed, two fundamental questions must bekept in mind:

How wi l l appl ied gene t ics be used?

Interest in the industrial use of biologicalprocesses stems from a merging of two paths:the revolution in scientific understanding of thenature of genetics; and the accelerated searchfor a sustainable society in which most indus-trial processes are based on the use of renew-able resources. The new genetic technologieswill spur that search in three ways: they willprovide a means of doing something biolog-ically—with renew able raw materials—that pre-viously required chemical processes using non-

renewable resources; they will offer more ef-ficient, more economical, less polluting ways forproducing both old and new p roducts; and theywill increase the yield of the plant and animalresources that are responsible for providing thewor ld’s sup plies of food, fibers, and som e fuels.

What a re the dan gers?

Even before scientists recognized the p oten-

tial power of app lied genetics, some qu estionedits consequences; for with its benefits, appearedhypothetical risks. Although most experts today

agree that the immediate hazards of the basicresearch itself appear to be minimal, nobodycan be certain about all the consequences of placing genetic characteristics in micro-orga-nisms, plants, and anima ls that have never car-ried them before. There are at least th ree sepa-rate areas of concern:

First, genetically engineered micro-organismsmight have potentially deleterious effects on hu-

man health, other living organisms, or the envi-ronment in general. Unlike toxic chemicals, or-ganisms may reproduce and spread of theirown accord ; if they are released into the env i -ronmen t, they may be imp ossible to control.

Second, some observers question whethersufficient know ledge exists to allow the extinc-tion of diverse species of “genetically inferior”plants and animals in favor of a few strains of “superior” ones. Evolution thus far has de-pend ed, in part, on genetic diversity; replacing

in natu re d iverse inferior strains by genetically

engineered superior strains may increase thesusceptibility of living things to disease and en-vironmental insults.

Finally, this new knowledge affects the un-derstand ing of life itself. It is tied to th e ultima te




questions of how humans view themselves and locating and monitoring its benefits and bur-what they legit imately cont ro l in the world . dens. That process requires knowledge. The fo l-

lowing sections of the report describe the im-Because of the significant and wide-ranging

scope of applied genetics, society as whole mustpacts of applied genetics on specific industries,

begin to debate the issues with a view toward al-and assess many of their consequences.



P a r t I

B i o t e c h n o l o g y

Chap ter 3-Gen etic Eng inee ring i n th e Ferment ation Techn ologies . . . . . . , . . . . . . . . . 49

Chapter 4-The Pharmaceutical Industry . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 59

Chapter 5-The Chemical Industry. . . . . . . . . . . . . , . . . . . . . . . . . . . . . . . . . , . . . . . . . 85

Chapter 6-The Food Processing Industry . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 107

(C hapter 7- The Use of Genetically Engineered Micro-Organisms in the Environment. . 117



Ch a p t e r 3

G e n e t i c E n g i n e e r i n g a n d t h e

F e r m e n t a t i o n T e c h n o l o g i e s



C h a p t e r 3

Page

Biotechn ology—An In trod uction . . . . . . . . . . . . . 49Fermentation . . . . . . . . . . . . . . . . . . . . . . . . . . . 49

Fermentation Industries . . . . . . . . . , . . . . . . . 50

Fermentation Using Wh ole Living Cells . . . . . . 51

The Process of Enzyme Technology. . . . . . . . . 53Comparative Advantages of Fermentations

Using w hole Cells an d Isolated Enzymes. . . . 54The Relationship of Genetics to Fermentation . 54Fermentation and Industry . . . . . . . . . . . . . , . 55

Table

Figures

Figure No.

16. Diagram of Produ cts Available From Cells . . . 5117. Features o f a Stan dard Ferment er . . . . . . . . . 5218. Immobilized Cell System . . . . . . . . . . . . . . . . 52

19. Diagram of Conversion of Raw Materialto Product. . . . . . . . . . . . . . . . . . . . . . . . . .

Table No. Page

2. Enzyme Products . . . . . . . . . . . . . . . . . . . . . . 54



C h a p t e r 3

G e n e t i c E n g i n e e r i n g a n d

t h e F e r m e n t a t i on T ec h n olo gie s

B i o t e c h n o l o g y — a n i n t r o d u c t i o n

Biotechnology involves the use in industry of living organisms or their components (such asenzymes). It includes the introduction of geneti-cally engineered micro-organisms into a varietyof industrial processes.

The ph armaceutical, chemical, and food proc-essing ind ustries, in that ord er, are most likelyto take advantage of advan ces in molecular ge-netics. Others that might also be affected, al-though not as immediately, are the mining,

crude oil recovery, and pollution control in-dustries.

Because nearly all the products of biotechnol-ogy are manufactured by micro-organisms, fer-mentation is an indispensable element of bio-technology’s support system. The pharmaceuti-cal industry, the earliest beneficiary of the newknowledge, is already producing pharmaceu-ticals derived from genetically engineeredmicro-organisms. The chemical industry willtake longer to make use of biotechnology, butthe ultimate impact may be enorm ous. The food

processing industry will probably be affectedlast.

This report examines many of the pharma-ceutical industry’s products in detail, as well as

F e r m e n t a t i o n

There are several ways tha t DNA can be cut,spliced, or otherwise altered. But engineered

DNA by itself is a static molecule. To be any-thing more than the end of a laboratory exer-

cise, the molecule must be integrated into a sys-tem of p rodu ction; to have an impact on societyat large, it must become a compon ent of an in-du strial or otherw ise useful p rocess.

The process that is central to the economic

some of the secondary impacts that the technol-ogies might have. Because the chemical andfood industries will feel the major impact of bio-technology later, specific impacts are less cer-tain and particular products are less identifi-able. The mining, oil recovery, and pollutioncontrol industries are also candidates for theuse of genetic technologies. How ever, becauseof technical, scientific, legal, and economic un-certainties, the success of applications in theseindu stries is more speculative.

The generalizations made with respect toeach of the indu stries shou ld be viewed as justthat—generalizations. Because a wid e array of products can be made biologically, and becausedifferent factors influence each instance of pro-duction, isolated examples of success may ap-pear throughout the industries at approximate-ly the same time. In almost every case, specificpred ictions can only be mad e on a prod uct-by-prod uct basis; for while it may be tru e that bio-technology’s overall impact will be profound,identifying many of the products most likely to

be affected remains speculative.

success of biotechnology has been around forcenturies. It is fermentation, essentially theprocess used to make wine and beer. It can alsoproduce organic chemical compounds using

micro-organisms or their enzymes.Over the years, the scope and efficiency of

the fermentation process has been grad ually im-proved and refined. Two processes now exist,both of which will benefit from genetic engi-

49




neering. In fermentation technology, living or-ganisms serve as miniature factories, convert-ing raw materials into end products. In enzymetechnology, biological catalysts extracted fromthose living organisms are used to make theproducts.

Fe rme n ta t ion indus t r i e s

The food processing, chemical, and p harm a-ceutical ind ustries are the th ree major users of fermentation today. The food ind ustry w as thefirst to exploit micro-organisms to producealcoholic beverages and fermented foods. Mid-16th century records describe highly sophisti-cated methods of fermentation technology. Heatprocessing techniques, for example, anticipatedpasteurization by several centuries.

[n the early 20th century, the chemical in-

du stry began to use the technology to prod uceorganic solvents like ethanol, and enzymes likeamylase, used at the t ime to treat t extiles. Thechemical industry’s interest in fermentationarose as the field of biochemistry took shapearound the turn of the century. But it was notuntil World War I that wartime needs for theorganic solvent acetone—to produce the cor-dite u sed in explosives–substantially increasedresearch into the potential of fermentation.Thirty years later after World War 11, the phar-maceutical industry followed the chemical in-dustry’s lead, applying fermentation to the pro-

du ction of vitamins and new antibiotics.

Today, approximately 200 companies in the

United States and over 500 worldwide usefermentation technologies to produce a widevariety of products. Most use them as part of prod uction processes, usually in food p rocess-ing. But others manufacture either proteins,wh ich can be considered primary prod ucts, or ahost of secondary products, which these pro-teins help produce. For genes can make en-zymes, which are proteins; and the enzymescan help make alcohol, methane, antibiotics,

and many other substances.

Proteins, the p rimary p rodu cts, function as:

enzymes such as asparaginase which areused in the treatment of leukemia;

q

q

q

q

structural components, such as collagen,used in skin transplants following burntrauma;certain hormones, such as insulin andhuman growth hormone;substances in the immune system, such as

antibodies and interferon; andspecialized functional components, such ashemoglobin.

Fermentation technologies are so useful for pro-ducing proteins partly because these are thedirect products of genes. But proteins (as en-zymes) can also be used in thousands of addi-tional conversions to produce practically anyorganic chemical and many inorganic ones aswell: (See figure 16. )

Figure 16.-Diagram of Products AvailableFrom Cells

isolated

B

Raw materialProduct

In (A) DNA directs the formation of a protein, such as in-sulin, which is itself the desired product. In (B), DNA directsthe formation of an enzyme which, in turn, converts someraw material, such as sugar, to a product, such as ethanol.




Ch. 3—Genetic Engineering and the Fermentation Technologies . 51

carbohydrates, such as fructose sweeten-ers;lipids, such as vitamins A, E, and K;alcohols, such as eth anol;other organic comp ound s, such as acetone;and

inorganic chemicals, such as ammonia, foruse in fertilizers.

Fermentation is not the only w ay to m anufac-ture or isolate these products. Some are tradi-tionally produ ced by other methods. If a changefrom one process to another is to occur, botheconomic and societal pressures w ill help d eter-mine wh ether an innovative approach w ill beused to produce a particular product. Alan Bullhas identified four stimuli for change and in-novation:

1

1. abundance of a potentially useful raw

material;2. scarcity of an established product;3. discovery of a new prod uct; and4. environmental concerns.

And conditions existing today have added a fifthstimulus:

5. scarcity of a currently used raw material.

Each of these factors h as tend ed to acceleratethe application of fermentation.

1. Abundance of a potentially useful raw ma-terial.—The use of a raw material can be

the d riving force in developing a pr ocess.When straight chain hydrocarbons (n-al-kanes) were produced on a large scale aspetroleum refinery byproducts, fermenta-tion processes were developed to convertthem to single-cell proteins for use in ani-mal feed.

2. Scarcity of an established product.—Thenew-found potential for producing humanhormones through fermentation technol-ogy is a major impetus to the industry to-day. Similarly, many organic compoundsonce obtained by other processes—like

citric acid, which was extracted directly

‘ / \ . ‘I”. lhJll, 11 . [:. F l lW r CM ) C i , and (:. Rat ledge, kficrobia/ Techndo,gv:

(~urrent Sfafe, Future Prospects, Z$lth Symposium ot ’ the Society for

(k l l f> l ’ i i l hlicrut)iub~y al 1 llll)f~rsitv uf (:iilllt)l’itl~f~, April 1979

((:illllt)l’idg(’, l’;ll~lillld: (kmt) r idge [~l;ilwrsit}~ Press , 1 979) , pp . 4 -8 .

3.

4.

5.

from citrus fruits—are now made by fer-mentation. As a result of more efficienttechnology, products from vitamin B12 tosteroids have come into wider use. Discovery of a new product.—The discovery

that antibiotics were produced by micro-

organisms sparked searches for an entirelynew group of products. Several thousand

antibiotics have been discovered to date, of

which over a hundred have proved to be

clinically useful.

Environmental concerns.—The problems of sewage treatment and the need for newsources of energy h ave triggered a searchfor method s to convert sewage and mu nici-pal w astes to methane, the principal com-ponent of natural gas. Because micro-orga-nisms play a major role in the natural cy-cling of organic compound s, fermentation

has been one method used for the conver-sion.Scarcity of a currently used raw materi-a/ .—Because the Earth’s sup plies of fossilfuels are rapidly d wind ling, there is intenseinterest in finding m ethods for convertingother raw materials to fuel. Fermentationoffers a major approach to such conver-sions.

Fermentation technologies can be effective ineach of these situations because of their out-standing versatility and relative simplicity. Theprocesses of fermentation are basically identi-

cal, no matter what organism is selected, what

medium used, or what product formed. The

same apparatus, with minor modifications, can

be used to produ ce a dru g, an agricultural prod -uct, a chemical, or an animal feed sup plement.

Ferm e n ta t ion us in g w h o le l i v ing c el l s

Originally, fermentation used some of themost primitive forms of plant life as cell fac-tories. Bacteria were u sed to m ake yogurt andantibiotics, yeasts to ferment wine, and thefilamentous fun gi or molds to produce organicacids. More recently, fermen tation t echnologyhas begun to use cells derived from higherplants and animals under growth conditionsknown as cell or tissue culture. In all cases,large quantities of cells with uniform character-



Ch. 3—Genetic Engineering and the Fermentation Technologies q 53

individual steps to complete the conversion. In achemical synthesis, the raw material (shown infigure 19 as a) might have to be transformed toan intermed iate b, wh ich, in turn, might have tobe converted to intermediates c and d beforefinal conversion to the product e–each step

necessitating the recovery of its products beforethe next conversion. In fermentation technol-ogy, all steps take place within those m iniaturechemical factories, the micro-organisms; themicrobial chemist merely adds the raw material

a and recovers the product e.

A wid e variety of carbohydrate raw materialscan be used in fermentation. These can be pu resubstances (sucrose or table sugar, glucose, orfructose) or complex mixtures still in theiroriginal form (cornstalks, potato mash, sugar-cane, sugar beets, orcellulose). They can be of

recent biological origin (biomass) or derivedfrom fossil fuels (methan e or oil). The ava ilabili-ty of raw materials varies from country to coun-

3) In the Chemical Conversion of raw material a to final

product e, intermediates b, c, and d must be synthe-

sized. Each intermediate must be recovered and purifiedbefore it can be used in the next step of the conversion.

)) A cell can perform the same conversion of a to e, butwith the advantage that the chemist does not have to

deal with the intermediates: the raw material a k simply

added and the final product e, recovered.

OURCE: Office of Technology Assessment.

try and even from region to region within acountry; the economics of the p rodu ction p roc-ess varies accord ingly.

The cost of the raw material can contributesignificantly to the cost of production. Usually,the most useful micro-organisms are those thatconsume readily available inexpensive raw ma-terials. For large volume, low-priced products(such as commodity chemicals), the relationshipbetween the cost of the raw material and thecost of the end product is significant. For lowvolume, high-priced products (such as certainpharmaceuticals), the relationship is negligible.

The process o f enzyme technology

Although live yeast had been u sed for severalthousand years in the production of fermentedfoods and beverages, it w as not u ntil 1878 thatthe active agents of the fermentation processwere given the name “enzymes” (from theGreek, meaning “in yeast”). The inanimatenature of enzymes was d emonstrated less thantwo d ecades later wh en it was shown that ex-tracts from yeast cells could effect the conver-sion of glucose to ethanol. Finally, their actualchemical nature was established in 1926 withthe purification and crystallization of theenzyme urease.

Fermentation carried out by live cells pro-vided the conceptual basis for designing fer-

mentation p rocesses based on isolated enzymes.A single enzyme situated within a living cell isneeded to convert a raw m aterial into a p rod-uct. A lactose-fermenting organism, e.g., can beused to convert the sugar lactose, which isfound in m ilk, to glucose (and galactose). But if the actual enzyme responsible for the conver-sion is identified, it can be extracted from thecell and used in place of a living cell. Thepurified enzyme carries out the same conver-sion as the cell, breaking d own the raw materialin the absence of any viable micro-organism. Anenzym e that acts inside a cell to convert a raw

material to a prod uct can also do this outside of the cell.

Both batch and continuous methods are usedin enzyme technology. However, in the batchmethod , the enzymes cannot be recovered eco-



54 . Impacts of Applied Genetics—Micro-Organisms, Plants, and Animals .

nomically, and new enzymes must be added foreach production cycle. Furthermore, the en-zymes are difficult to separate from the endproduct and constitute a potential contaminant.Because enzymes used in the continuous meth-od are reusable and tend not to be found in the

produ ct, the continuous method is the methodof choice for m ost p rocesses. Depend ing on thedesired conversion, the immobilized micro-organisms of figure 18 could be replaced by anappropriate immobilized enzyme.

Although more than 2,000 enzymes havebeen d iscovered, fewer than 50 are currently of industrial importance. Nevertheless, two majorfeatures of enzymes make them so desirable:their specificity and their ability to operateunder relatively mild conditions of temperatureand pressure. (The most frequently used. en-

zymes are listed in table 2.)

C ompara t i v e adv an tage s o f

f e rme n ta t ions us ing w ho le c e l l s

and i so la te d e nzy me s

At present, it is still uncertain whether theuse of whole cells or isolated enzymes will bemore useful in the long run. There are advan-tages and disadvan tages to each. The role of ge-netic engineering in the future of the industry,

Table 2.—Enzyme Products

Commercially Currentavailable before: production

Source/name 1900 1950 1980 tons/yr

Animal Rennet. . . . . . . . . . . . . . . X 2Trypsin. . . . . . . . . . . . . . . x 15Pepsin . . . . . . . . . . . . . . . x 5

PlantMalt amylase. . . . . . . . . . X 10,000Papain . . . . . . . . . . . . . . . x 100Microbial Koji. . . . . . . . . . . . . . . . . . XFungal protease . . . . . . . X 10Bacillus protease . . . . . . x 500

Amyloglucosidase . . . . . x 300Fungal amylase. . . . . . . . X 10Bacterial amylase. . . . . . x 300Pectinase. . . . . . . . . . . . . x 10Glucose isomerase. . . . . x 50Microbial rennet . . . . . . . x 10


however, will be partly determined by whichmethod is chosen. With isolated enzymes, ge-netic manipulation can read ily increase the sup -ply of enzym es, wh ile with w hole organisms, awide variety of manipulations is possible in con-structing m ore prod uctive strains.

T h e re la t ionsh ip o f ge ne t i cs

to f e rme n ta t ion

Applied genetics is intimately tied to fermen-tation technology, since finding a suitable spe-cies of micro-organism is u sually the first step indeveloping a fermentation technique. Until re-cently, geneticists have had to search for anorganism that already produced the neededprodu ct. However, through genetic manipula-tion a totally new capability can be engineered;

micro-organisms can be made to produce sub-stances beyond their natural capacities. Themost striking successes have been in the phar-maceutical industry, where hu man genes havebeen transferred to bacteria to prod uce insulin,growth hormone, interferon, thymosin a-1, andsomatostatin, (See ch. 4.)

In general, once a species is found, conven-tional methods have been used to induce muta-tions that can produce even more of the desiredcompound. The geneticist searches from amonghundreds of mutants for the one micro-orga-

nism that produces most efficiently. Most of theman y method s at the m icrobiologist’s disposalinvolve trial-and-error. Newer genetic tech.nologies, such as the u se of recombinant DN A(rDNA), allow ap proaches in w hich useful genetic traits can be inserted directly into the m icroorganism.

The curren t indu strial app roach to fermentation technologies therefore considers two problems: First, whether a biological process carproduce a particular product; and second, whamicro-organism has the greatest potential for

prod uction and how the desired characteristic;can be engineered for it. Finding the desiredmicro-organism and impr oving its capability iso fundamental to the fermentation industrythat geneticists have become important members of fermentation research teams.



Ch. 3—Genetic Engineering and the Fermentation Technologies q 55

Genetic engineering can increase an orga-nism’s p rodu ctive capability (a change th at canmake a p rocess economically comp etitive); but itcan also be used to constru ct strains w ith char-acteristics other than higher productivity. Prop-erties such as objectionable color, odor, or slime

can be removed. The formation of spores thatcould lead to airborne spread of the micro-organism can be suppressed. The formation of harmful byproducts can be eliminated or re-duced. Other properties, such as resistance tobacterial viruses and increased genetic stability,can be given to micro-organisms that lack them.

Applying recent genetic engineering tech-niques to the prod uction of ind ustrially valuableenzymes may also prove useful in the future.For example, a strain of micro-organism thatcarries the genes for a desired enzym e may be

path ogenic. If the genes that expr ess (prod uce)the enzyme can be transferred to an innocuousmicro-organism, the enzyme can be producedsafely.

CURRENT TECHNICAL LIMITS ONGENETIC ENGINEERING

Despite the many genetic manipulations thatare theoretically possible, there are severalnotable technical limitations:

q

q

q

Genetic maps—the identification of the lo-cation of desired genes on various chromo-somes have not been constructed for most

industrially useful micro-organisms.Genetic systems for industrially usefulmicro-organisms, such as the availability of useful vectors, are at an early stage of development.Physiological pathways—the sequence of enzym atic steps leading from a raw m ate-rial to the d esired p roduct, are not knownfor many chemicals. Much basic researchwill be necessary to identify all the steps.The number of genes necessary for the con-version is a major limitation. Currently,rDNA is most useful when only a singleeasily identifiable gene is need ed. It is moredifficult to u se wh en several genes must betransferred. Finally, the problems are for-midable, if not impossible, when the geneshave not yet been identified. This is the

case with many traits of agronomic impor-tance, such as p lant height.Even if the genes are identified and suc-cessfully transferred, methods must be de-veloped to recognize the bacteria that re-ceived th em. Therefore, the need to d evel-

op appropriate selection methods has im-peded the application of molecular ge-netics.

As a consequence of these limitations, geneticengineering will be applied to the developmentof capabilities that require th e transfer of onlyone or a few identified genes.

F e r m e n t a t i o n a n d i n d u s t r y

Genetic engineering is not in itself an ind us-try, but a technology used at the laboratorylevel. It allows the researcher to alter the hered-itary apparatus of a living cell so that the cellcan produce more or different chemicals, orperform completely new functions. The alteredcell, or more appropriately the population of altered identical cells is, in tu rn, u sed in indu s-trial production. It is within this framework thatthe impacts of applied genetics in the various in-dustries is examined.

Regardless of the industry, the same threecriteria must be met before genetic technologiescan become commercially feasible. These cri-teria represent major constraints that industry

mu st overcome before genetic engineering canplay a part in bringing a product to market.They include th e need for:

1. a u seful biochemical prod uct;2. a useful biological fermentation ap proach

to commercial produ ction; and3. a useful genetic approach to increase the

efficiency of production.

The three criteria interrelate and can be m etin any order; the demonstration of usefulnesscan begin with any of the three. Insulin, e.g.,was first found to have value in therapy;fermentation was then shown to be useful inits production; and, now genetic engineeringprom ises to m ake the fermentation p rocess eco-nom ically competitive. In contrast, the value of thymosin a-1, has not yet been proved, although



. Impacts of Applied Genet ics—Micro-Organisms, Plants, and Animals

the usefulness of genetic engineering and fer- fulness of genetic technologies must be proved.mentation in its production have been demon- In others, the genetic technologies make pro-strated. duction at the industrial level possible, but their

As these examples indicate, the limits on amarket has not yet been established. In still

product’s commercial potential vary with theothers, the feasibility of fermentation is the ma-

product. In some cases, the usefulness of the jor problem.

produ ct has already been shown, and th e use-



Ch a p t e r 4

Th e P h a r m a c eu t ica l In d u s t r y



Ch a p t e r 4

Page

Backgrou nd . . . . . . . . . . . . . . . . . . . . . . . . . . . . 59Past Uses of Genetics. . . . . . . . . . . . . . . . . . . . 59

Potential Uses of Molecular Genetic Technologies 61

Hormones . . . . . . . . . . . . . . . . . . . . . . . . . . . . 61

Insulin . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 65Grow th Horm one . . . . . . . . . . . . . . . . . . . . . 67Other Hormones. . . . . . . . . . . . . . . . . . . . . . 67

Immunoproteins . . . . . . . . . . . . . . . . . . . . . . . 68Antigens (Vaccines). . . . . . . . . . . . . . . . . . . . 68Interferon . . . . . . . . . .. . . $ . . . . . . . . . . . . 70Lymphokines and Cytokines . . . . . . . . . . . . . 71Antibodies . . . . . . . . . . . . . . . . . . . . . . . . . . 71

Enzymes and Other Proteins . . . . . . . . . . . . . . 72Enzymes . . . . . . . . . . . . . . . . . . . . . . . . . . . . 72Other Protein s . . . . . . . . . . . . . . . . . . . . . . . 74

Antibiotics . . . . . . . . . . . . . . . . . . . . . . . . . . . . 75Non protein Pha rmaceuticals . . . . . . . . . . . . . . 76

Impacts . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 77

Technical N otes . . . . . . . . . . . . . . . . . . . . . . . . . 80

Tables

Table No. Page

3. Large Human Polypeptides PotentiallyAttractive for Biosynthesis . . . . . . . . . . . . . . . 61

Table No. Page

4. Naturally Occurring Small Peptides of Poten tial Medical In terest . . . . . . . . . . . . . . . . 64

5, Summary of Potential Methods forInterferon Production. . . . . . . . . . . . . . . . . . . 70

6. Immunoassays . . . . . . . . . . . . . . . . . . . . . . . . 737. Diseases Amenable to Dru gs Produced by

Genetic Engineering in the Ph armaceuticalIndustry . . . . . . . . . . . . . . . . . . . . . . . . . . . . 78

8. Major Diseases for Which Vaccines Need ToBe Developed . . . . . . . . . . . . . . . . . . . . . . . . . 78

Figures

.Figure No. Page

20. The Development of a High Penicillin-Producing Strain via Genetic Manipulation . . 60

21. The Product Development Process forGenetically Engineered Pharmaceuticals . . . . 63

22. The Amin o Acid Sequ ence of Proinsulin . . . . 6523. Recombinant DNA Strategy for Making

Foot-and-Mouth Disease Vaccine . . . . . . . . . . 69



Ch a p t e r 4

T h e P h a r m a c e u t i c a l I n d u s t r y,

B a c k g r o u n d

The domestic sales of prescription drugs by

U.S. pharmaceutical companies exceeded $7.5billion in 1979. Of these, app roximately 20 per -cent were products for which fermentationprocesses played a significant role. They in-clud ed anti-infective agents, vitamins, and bio-logical, such as vaccines and horm ones. Genet-ics is expected to be particularly useful in theproduction of these pharmaceuticals and bio-logical) which can on ly be obtained by extrac-tion from hu man or animal tissues and fluids.

Although the pharmaceutical industry wasthe last to adopt traditional fermentation tech-nologies, it has been the first indu stry to m akewid espread u se of such adv anced genetic tech-nologies as recombinan t DN A (rDNA) and cellfusion. Two major factors triggered the use of genetics in the phar maceutical indu stry:

q

q

The biological sources of many ph armaco-logically active products are micro-orga-nisms, which are readily amenable to ge-netic engineering.The major advances in molecular geneticengineering have been mad e und er an in-stitutional structure that allocates fundslargely to biom edical research. H ence, the

Federal support system has tended to fos-ter stud ies that have a s their ostensible goalthe improvem ent of health.

Two factors, however, have tended to dis-courage the application of genetics in the chem-cal and food indu stries. In th e former, econom-c considerations have not allowed biologicalprod uction systems to be competitive with theexisting forms of chemical conversion, withrare exceptions. And in the latter, social and in-stitutional considerations have not favored thedevelopment of foods to which genetic engi-

neering might make a contribution.

Past uses o f gene t ics

Genetic manipulation of biological systemsfor the production of pharmaceuticals has twogeneral goals:

1. to increase the level or efficiency of theproduction of pharmaceuticals with prov-en or p otential value; and

2. to produce totally new pharmaceuticalsand compound s not found in nature.

The first goal has h ad the strongest influenceon the industry. It has been almost axiomaticthat if a naturally occurring organism can pro-duce a pharmacologically valuable substance,genetic manipulation can increase the output.The

q

q

q

following are three classic examples.

The genetic improvement of penicillin pro-du ction is an example of the elaborate long-term efforts that can lead to dramaticincreases. The original strains of Penicilli-um chrysogenum, NRRL-1951 ) were treatedwith chemicals and irradiation throughsuccessive stages, as shown in figure 20,

un til the strain E-15.1 was d eveloped. Thisstrain had a 55-fold improvement in pro-ductivity over the fungus in which penicil-lin w as originally recognized–the Flemingstrain.

Chemically induced mutations improved astrain of Escherichia coli to the point whereit prod uced over 100 times m ore L-asparag-inase (which is used to fight leukemia) thanthe original strain. This increase made thetask of isolating and purifying the pharma-ceutical much easier, and resulted in low -ering the cost of a cour se of therapy fromnear ly $15,000 to ap proximately $300.

Genetic manipulation sufficiently improved

the production of the antibiotic, gentami-

59



Ch. 4—The Pharmaceutical Industry q 61

P o t e n t i a l u se s o f m o l e c u l a r g e n e t i c t e c h n o l o g i e s —

Polypeptides—proteins—are the first abun-dant end products of genes. They include pep-tide hormones, enzymes, antibodies, and cer-

tain vaccines. Producing them is the goal of most current efforts to harness geneticallydirected p rocesses. However, it is just a matterof time and the evolution of technology beforecomplex nonproteins like antibiotics can also bemanufactured through rDNA techniques.

H o r m o n e s

The most advanced applications of geneticstoday , in term s of technological soph isticationand commercial developm ent, are in the field of hormones, the potent messenger molecules that

help the body coordinate the actions of varioustissues. (See Tech. Note 1, p. 80.) The capacity tosynthesize proteins through genetic engineer-ing has stemmed in large part from attemp ts toprepare hu man peptide h ormones (like insulinand growth hormone). The diseases caused bytheir d eficiencies are p resently treated w ith ex-tracts made from animal or human glands.

The merits of engineering other peptide hor-mones depend on understanding their actionsand those of their derivatives and analogs.Evidence that they might be used to improve

the treatment of diabetes, to promote woundhealing, or to stimu late the regrowth of nerveswill stimulate new scientific investigations.Other relatively small polypeptides that influ-ence the sensation of p ain, appetite supp ression,and cognition and memory enhancement arealso being tested. If they prove useful, they willunquestionably be evaluated for production viafermentation.

While certain hormones have already at-tained a place in pharmacology, their testingand use has been hindered to some extent bytheir scarcity and high cost. Until recently,animal glands, human-cadaver glands, andurine were the only sources from which theycould be drawn. Their use is also limitedbecause polypeptide hormones must be ad-ministered by injection. They are digested if

they are taken orally, a process that curtailstheir usefulness and causes side-effects.

There are four technologies for producingpolypeptide hormones and polypeptides:

. extraction from human or animal organs,

serum, or urine;q chemical synthesis;

q production by cells in tissue culture; andq prod uction by m icrobial fermentation after

genetic engineering.

One m ajor factor in d eciding w hich technol-ogy is best for which hormone is the length of the hormone’s amino acid chains. (See table 3.)Modern methods of chemical synthesis have

made the preparation of low-molecular weightpolypeptides a fairly straightforward t ask, andchemically synthesized hormones up to at least32 amino acids (AA) in length—like calcitonin

Table 3.—Large Human Polypeptides PotentiallyAttractive for Biosynthesis

Amino acid Molecularresidues weight

Proactin . . . . . . . . . . . . . . . . . . . . .Placental Iactogen . . . . . . . . . . . . .q Growth hormone. . . . . . . . . . . . . .Nerve growth factor . . . . . . . . . . . .Parathyroid hormone (PTH). . . . . .Prokm.din . . . . . . . . . . . . . . . . . . . .

Insulin-like growth factors(IGF-I & IGF-2) . . . . . . . . . . . . . . .

Epidermal growth factor . . . . . . . .q insulin . . . . . . . . . . . . . . . . . . . . . .Thymopoietin . . . . . . . . . . . . . . . . .Gastric inhibitory polypeptide

(GIP) . . . . . . . . . . . . . . . . . . . . . . .q Corticotropin (ACTH) . . . . . . . . . .Cholecystokinin (CCK-39) . . . . . . .Big gastrin (BG).. . . . . . . . . . . . . . .Active fragment of PTH . . . . . . . . .Cholecystokinin (CCK-33) . . . . . . .q Calcitonin . . . . . . . . . . . . . . . . . . .

Endorphins . . . . . . . . . . . . . . . . . . .q Glucagon . . . . . . . . . . . . . . . . . . . .Thymosin-a1 . . . . . . . . . . . . . . . . . .

Vasoactive intestinal peptide (VIP)q Secretin. . . . . . . . . . . . . . . . . . . . .q Active fragment of ACTH. . . . . . .Motilin . . . . . . . . . . . . . . . . . . . . . . .

q Currently used in medical practice.


1981921911188482

70,67

5149

43393934343332

312928

28272422

22,005 13,000 9,562 bovine

7,649,74716,1005,734

5,104 porcine4,567 porcine

4,109 bovine3,918 porcine3,421 human3,435 salmon3,4653,483 porcine3,108

3,326 porcine

2,698



62 . Impacts o/Applied Genetics—Micro-Organisms, Plants, and Animals

–have become competitive with those derivedfrom current biological sources. Since frag-ments of peptide hormones often express activ-ities comparable or sometimes superior to theintact hormone, a significant advantage of chemical synthesis for research purposes is that

analogs having slight pharmacological differ-ences from natural hormones can be preparedby incorporating different amino acids intotheir structures. In principle however, geneti-cally engineered biosynthetic schemes can bedevised for most desirable peptide hormonesand their analogs, although the practicality of doing so must be assessed on a case-by-casebasis. Ultimately, the principal factors bearingon the practicality of the competing alternativesare:

q

q

q

q

The cost of raw materials. For genetically

engineered biosynthesis, this includes thecost of the nu trient broth plu s some amor-tization of the cost of developing the syn-thetic organism. In the case of chemicalsynthesis, it includes the cost of the pureamino acid subunits plus the chemicalsused as activating, protecting, coupling, lib-erating, and supporting agents in the proc-ess.The different costs of separating the de-sired product from the cellular debris andthe culture medium in biological produc-tion, and from the supporting resin, by-

prod ucts, and excess reagents in chemicalsynthesis.The cost of pu rification and freedom fromtoxic contaminants. The process is moreexpensive for biologically produced materi-al than for materials produced by conven-tional chemistry, although h ormon es fromany source can be contaminated .Differences in the costs of labor and equip-ment. Chemical synthesis involves a se-quence of similar (but different) operationsduring a time period roughly proportionalto the length of the amino acid chain (three

AA per d ay) in an ap paratus large enoughto produce 100 grams (g) to 1 kilogram (kg)per batch; biological fermentations use vats—with capacities of several thousand gal-lons–for a few days, regardless of thelength of the amino acid chain.

. The cost and suitability of comparablematerials gathered from organs or fluidsobtained from animals or p eople.

In the past decade, some simpler hormoneshave been chemically synthesized and a few are

being marketed . However, synthesizing glyco-proteins—proteins bound to carbohydrates—isstill beyond the capabilities of chemists. Dataobtained from companies directly involved inthe pr odu ction of peptides by chemical synthe-sis indicate that the cost of chemically preparingpolypeptides of up to 50 AA in length is ex-tremely sensitive to volume (see Tech. Note 2, p.80,); although the costs are high, the productionof large quantities by chemical synthesis offersa competitive produ ction method .

Nevertheless, rDNA production, also knownas molecular cloning, has already been u sed toproduce low-molecular weight polypeptides. In1977, researchers at Genentech, Inc., a smallbiotechnology company in California, inserted atotally synthetic DNA sequence into an E. coliplasmid and demonstrated that it led to the pro-du ction of the 14 AA p olypeptide sequence cor-respond ing to somatostatin, a hormone found inthe brain. The knowledge of somatostatin’samino acid sequence made the experiment pos-sible, and the existence of sensitive assays al-lowed the hormone’s expression to be detected.Although the primary motive for using this par-

ticular hormone for the first demonstration wassimply to show that it could be don e, Genentechhas announced that it plans to market itsgenetically engineered molecule for researchpu rposes. (See figure 21.)

Somatostatin is one of about 20 recognizedsmall human polypeptides that can be madewithout difficulty by chemical synthesis. (Seetable 4.) Unless a sizable market is found for oneof them, it is unlikely that fermentation methods will be developed in the foreseeable futureSome small peptides that may justify the development of a biosynthetic process of productionare:

q The seven AA sequence known as MSHACTH 4-10, which is repu ted to influencemem ory, concentration, and other p sychelogical-behavioral effects: should such




Figure 21.—The Product Development Process for Genetically Engineered Pharmaceuticals

Micro-organisms such as E. coli

The development process begins by obtaining DNA either through organic synthesis (1) or derived from biological sources such as tissues(2). The DNA obtained from one or both sources is tailored to form the basic “gene” (3) which contains the genetic information to “code” for adesired product, such as human interferon or human insulin. Control signals(4) containing plasmids (6) are isolated from micro-organisms suchas E. coli; cut open (7) and spliced back (8) together with genes and control signals to form “recombinant DNA” molecules. These molecules arethen introduced into a host cell (9).

Each plasm id is copied many times in a cell (10). Each cell then translates the information contained in these plasmids into the desired pro-duct, a process called “expression” (11). Cells divide (12) and pass on to their offspring the same genetic information contained in the parentcell.

Fermentation of large populations of genetically engineered micro-organisms is first done in shaker flasks (13), and then in small fermenters(14) to determine growth conditions, and eventually in larger fermentation tanks (15). Cellular extract obtained from the fermentation process isthen separated, purified (16), and packaged (17) for health care applications.

Health care products are first tested in animal studies (18) to demonstrate a product’s pharmacological activity and safety, In the UnitedStates, an investigational new drug application (19) is submitted to begin human clinical trials to establish safety and efficacy. Followingclinical testing (20), a new drug application (N DA) (21) is filed with the Food and Drug Administration (FDA). When the NDA has been reviewedand approved by the FDA the product maybe marketed in the United States (22).

SOURCE: Genentech, Inc.



64 . Impacts of Applied Genetics—Micro-0rganisms,Plants, and Animals

Table 4.-Naturally Occurring Small Peptides ofPotential Medical interest

Number of Molecularamino acids weight

Dynorphin . . . . . . . . . . . . . . . . . . . . 17Little gastrin (LG) . . . . . . . . . . . . . . 17Somatostatin. . . . . . . . . . . . . . . . . . 14

Bombesin . . . . . . . . . . . . . . . . . . . . 14Melanocyte stimulating hormone. 13Active dynorphin fragment . . . . . . 13Neurotensin . . . . . . . . . . . . . . . . . . 13Mini-gastrin (G13) . . . . . . . . . . . . . . 13Substance? . . . . . . . . . . . . . . . . . . . 11Luteinizing hormone-releasing

hormone(LNRH). . . . . . . . . . . . . 10Active fragment of CCK. . . . . . . . . 10Angiotensin l . . . . . . . . . . . . . . . . . . 10Caerulein . . . . . . . . . . . . . . . . . . . . . 10Bradykinin . . . . . . . . . . . . . . . . . . . . 9q Vasopressin(ADH) . . . . . . . . . . . . 9q Oxytocin . . . . . . . . . . . . . . . . . . . . 9Facteur thymique serique(FTH).. 9Substance P(4-11)octapeptide. . . 8Angiotensin lI . . . . . . . . . . . . . . . . . 8

Angiotensin Ill. . . . . . . . . . . . . . . . . 7MSH/ACTH4-10 . . . . . . . . . . . . . . . 7Enkephalins. . . . . . . . . . . . . . . . . . . 5Active fragment of thymopoietin

(TP5). . . . . . . . . . . . . . . . . . . . . . . 5q Thyrotropin releasing hormone

(TRY) . . . . . . . . . . . . . . . . . . . . . . 3

2,1781,639

1,6201,655

l,347bovine

1,297l,252porcine1,060

1,007

1,046

931

575

362

.Currently used in medical practice.

SOURCE: Office of Technology Assessment

agents prove of value in w ider testing, theyhave an enorm ous potential for use.Both cholecystokinin (33 AA) and bombesin(10 AA), which have been shown to sup-press appetite, presumably as a satietysignal from stomach to brain: there is alarge market for antiobesity agents–ap-proximately $85 million per year at themanufacturer’s level.Several hormones, such as somatostatin,which are released by nerves in the hypo-thalamus of the brain to stimulate or in-hibit release of horm ones by the p ituitarygland: hormones produced by these glandsare crucial in human fertility; analogs of some are being investigated as possible

contraceptives.Calcitonin (32 AA), which is currently thelargest polypeptide prod uced by chemicalsynthesis for commercial pharmaceuticaluse: it is useful for pathologic bone dis-orders, such as Paget’s disease, that affect

up to 3 percent of the population over 40years of age, in WesternEurope.

. Adrenocorticotropic hormone (ACTH) (39AA), which promotes and maintains the

normal growth and development of the

adrenalglands and stimulates the secretion

of other hormones: in the United States,ACTH is used primarily as a diagnostic

agen t fo r ad rena l i nsu f f i c i ency , bu t i nprinciple, ACTH might be used for at least

one-third of the medical indications—like

rheumatic disorders, allergic states, and

eye inflammation—for which about 5 mil-

lion Americans annually receive corticos-

teroids.

Within the last 5 years, other small polypep-

tides have been identified in many tissues and

have been linked to a variety of activities. Some

certainly bind to the same receptor sites as the

pain-relieving opiates related to the morphinefamily. These peptides are called endogenousopiates: the smaller (5 AA) peptides are called

enkephalins and the larger (3 AA), endorphins.

Certain enkephalins produce brief analgesia

when injected directly into the brains of mice.

Synthetic analogs that are less susceptible to en-

zymatic inactivation produce longer analgesia

even if they are injected intravenously, as does

the larger endorphin molecule. Very recently,

a 17 AA polypeptide, dynorphin, was reportedto be the most p otent pain k iller yet found—it is

1,200 times more powerful than morphine.

The preparation of new analgesic agents ap-

pears a likely outcome of the new research, but

problems similar to those associated with clas-

sical opiates must be overcome. Consequently,

unnatural analogs—including some made with

amino acids not found in micro-organisms—

might prove more useful. The value of microbi-

al biosynthesis for these substances is ques-

tionable at this time. However, the importance

o f g e n e t i c technologies i n c l a r i f y i n g t h e

underlying mechanisms should not be under-

estimated.

Higher molecular weight polypeptides cannot

be made practically by chemical synthesis, and

must be extracted from human or animal tis-

sues or produced in cells growing in culture.




Now they can also be manufactured by fermen-tation using genetically designed bacteria, ashas been demonstrated by the production of in-sulin and hum an growth hormone.

INSULIN

Insulin, is composed of tw o chains—A and B—of amino acids. It is initially produced as asingle, long chain called p re-proinsu lin, which iscut into a shorter chain, proinsu lin. Proinsulin,in turn, is cut into the A and B chains when apiece is cleaved from th e midd le. (See figure 22.)Work on th e genetic engineering of insu lin has

proceeded quickly. A year after one group re-ported th at the insulin gene had been incorpo-rated into E. coli withou t expression, a secondgroup m anaged to grow colonies of E. coli thatactually excreted rat p roinsulin. Then, within acouple of months, workers at Genentech, in col-laboration w ith a group at City of Hop e MedicalCenter, announced the separate synthesis of theA (21 AA) and B (30 AA) chains of human insulin.The synthesis of the DNA sequences depend edon ad vances in organic chemistry as w ell as ingenetics. Six months were required simply tosynthesize the necessary building blocks.

Figure 22.—The Amino Acid Sequence of Proinsulin

Connecting peptide

N H

+

3

B chain

Proinsulin is composed of 84 amino acid residues. When the connecting peptide is removed,

the retaining A and B chains form the insulin molecule. The A chain contains 21 amino acids;the B chain contains 30 amino acids.





A comparison with the traditional source of animal insulin is interesting. If 0.5 milligram(mg) of pure insulin can be obtained from a literof fermentation brew, 2,000 liters (1) (roughly500 gal) would yield 1 g of purified insulin—theamount produced by about 16 lb of animal pan-

creas. If, on the other hand, the efficiency of prod uction could be increased to that achievedfor asparaginase (which is produced commer-cially by the same organism, E. coli), 2,000would yield 100 g of purified insulin—theamount extracted from 1,600 lb of pancreas.(The average diabetic uses the equivalent of about 2 mg of animal insulin per day.)

The extent of the actual demand for insulin isa controversial issue. Eli Lilly & Co. estimatesthat th ere are 60 million d iabetics in th e world(35 million in un derd eveloped countries, where

few are diagnosed or treated). Of the 25 millionin the d eveloped coun tries, perhap s 15 millionhave been diagnosed; according to Lilly’s esti-mate, 5 million are treated with insulin. Onlyone-fourth of those diabetics treated with in-sulin live in the United States, but they use 40 to50 percent of the insulin consumed in theworld . A nu mber of studies indicate that w hilethe emphasis on diet (alone) and oral antidia-betic dru gs varies, appr oximately 40 percent of American patients in large diabetes clinics orpractices take insulin injections. In the UnitedStates, diabetes ranks as the fifth most common

cause of death and second most common causeof blindness. Roughly 2 million persons requiredaily injections of insulin.

Today, at least, there is no real shortage of glands from slaughter houses for the produc-tion of animal (principally bovine and porcine)insulin. A stud y condu cted by the National Dia-betes Advisory Board (NDAB) concluded that amaximum demand and a minimum supplywou ld lead to shortages in th e 1990’s. Eli Lilly’sprojection, presented in that report, also antici-pates these shortages. But, N ovo Indu stri, a m a-

jor wor ld sup plier of insu lin, told the N DAB thatit estimates that th e 1976 free-world consu mp -tion of insulin of 51 X 10

9un its constituted on ly

23 percent of the potential supply, and the87X 10

9units projected for 1996 would only

equal 40 percent of the supply, assuming thatthe animal population stays constant.

For insulin, therefore, the limitation on bring-ing the fruits of genetic engineering to themarketplace is not technological but institu-tional. The drug must first be approved by theFood and Drug Administration (FDA) and thenmarketed as a product as good as or better than

the insulin extracted by conventional means.Lilly has stated that it anticipates a 6-monthtesting period in humans. Undoubtedly, FDAwill examine the evidence presented in th e in-vestigational new dru g ap plication (INDA) w ithspecial care. Its review will establish criteriathat may influence the review of subsequent ap -plications in at least the following requirements:

evidence that the amino acid sequence of the m aterial is identical to that of the nor-mal human hormone;freedom from bacterial endotoxins that

may cause fever at extremely low concen-trations—an inherent hazard associatedwith any process using E. coli; an dfreedom from byproducts, including sub-stances of very similar structure that maygive rise to rare acute or chron ic reactionsof the immu ne system.

Furthermore, as development continues, FDAmight require strict assurances that the mole-cules produced from batch to batch are not subject to subtle variations resulting from theirgenetic origin.

If the insulin obtained from rDN A techniquesmanages to pass FDA requirements, it mustovercome a second obstacle—competition in themarketplace. The clinical rationale for usinghum an rather than animal insulin rests on thedifferences in structure among insulins pro-duced by different species. Human and porcineinsulins for example, differ in a single aminoacid, while human and cattle insulins differwith respect to three. As far as is know n, thesevariations do not impair the effectiveness of theinsulin, but n o one has ever been in a position toconduct a significant test of the use of human

insulin in a diabetic population. Many conse-quences of the disease, such as retinopathy (ret-inal disease) and nephropathy (kidney disease),are not p revented by routine injection of animalinsulin. Patients also occasionally respond ad-versely or produ ce antibodies to animal insulin,with subsequent allergic or resistant reaction.



Ch. 4—The Pharmaceutical Industry . 67

It remains to be seen how many patients willbe better off with hu man insulin. The proof thatit improves therap y w ill take years. Progress onthe etiology of the disease—especially in identi-fying it in those at risk or in improving thedosage form and administration of insulin–may

have far more significant effects than new de-velopments in insu lin p rodu ction. Nevertheless,as long as p rivate enterprise sees fit to invest insuch developments, and as long as the cost of treating diabetics who respond properly to ani-mal insu lin is not increased, biological produc-tion of human insulin may become a kind of in-surance for diabetics within the next fewdecades.

GROWTH H ORMONE

The second polypeptide hormone currently acandidate for FDA approval is growth hormone

(GH). It is one of a family of closely related, rel-atively large pituitary peptide hormones—sin-gle-chain polyp eptid es 191- to 198-AA in length .It is best known for the growth it induces inmany soft tissues, cartilage, and bone, and it is arequirement for postnatal growth in man.

The growth of an organism is a highly com-plex process that depends on the correct bal-ance of many variables: The action of GH in thebody for example, depends on the presence of insulin, whose secretion is stimulated by GH.Und er some circumstan ces, one or more inter-mediary polypeptides produced under the in-

fluence of GH by the liver (and possibly thekidneys) may actua lly be the p roximate causesof some of the effects attributed to GH . In anycase, the biological significance of GH is mostclearly illustrated by the growth retardationthat characterizes its absence before puberty,and by the benefits of replacement therap y.

In the United States, most of the dem and forhuman growth hormone (hGH) is met by the Na-tional Pituitary Agency, which was created inthe early 1960’s by the College of Pathologistsand the National Institute of Arthritis, Metab-

olism, and Digestive Diseases (NIAMDD) to col-lect pituitary gland s from coroners and p rivatedonors. Under the p rograms of the NIAMDD,hGH is provided without charge to treat chil-dren with hypopituitarism, or dwarfism (about

1,600 patients, each of w hom receives therap yfor several years), and for research.

While the National Pituitary Agency feels thatit can satisfy the current d emand for hGH (seeTech. Note 3, p. 80.), it welcomes the promise of add itional hGH at relatively low cost to satisfyareas of research that are handicapped more bya scarcity of funds than by a scarcity of the hor-mone. However, if hGH is shown to be thera-peutically valuable in these areas, widespreaduse could severely strain the present supply. Atpresent, the potential seems greatest for pa-tients with:

q

q

q

q

q

senile osteoporosis (bone decalcification);other nonpituitary growth deficiencies suchas Turner’s syndrome (1 in 3,000 livefemale births);intrauterine growth retardation;

bleeding ulcers that cannot be controlledby other mean s; andburn, wound, and bone-fracture healing

Two groups have already announced theprep aration of micro-organisms w ith the capaci-ty for syn thesizing GH . (See Tech. Note 4, p. 80.)In December 1979, one of these groups—Genen-tech —requested and received permission fromthe N ational Institutes of Health (NIH), on therecommendation of the Recombinant DNA Ad-visory Committee (RAC), to scale-up its process.Its formation of a joint-venture w ith Kabi Gen

AB is typical of the kind of alliance that d evelopsas a result of the different expertise of groups inthe multidisciplinary biomedical field. Kabi hasbeen granted a New Drug Ap plication (NDA)und er which to market pituitary GH importedfrom abroad.

OTHER HORMONES

Additional polypeptide hormones targetedfor molecular cloning (rDNA production) in-clude:

q

q

Parathyroid horm one (84 AA), which may

be useful alone or in combination with cal-citonin for bone disorders such as osteo-porosis.Nerve growth factor (118 AA), which influ-ences the development, maintenance, and




repair of nerve cells and thu s could be sig-nificant for nerve restoration in surgery.

q Erythropoietin, a glycopeptide that is large-ly responsible for the regulation of bloodcell development. Its therapeutic applica-tions may range from hemorrhages and

burns to anemias and other hematologicconditions. (See Tech. Note 5, p. 80.)

I m m u n o p r o t e i n s

Immunoproteins include all the proteins thatare part of the immune system—antigens, inter-feron, cytokines, and antibodies. Since poly-pep tides, the primary pr odu cts of every molec-ular cloning scheme, are at the heart of immu-nology, developments m ade p ossible by recentbreakthroughs will presumably affect the entirefield. There is little doubt that a pp lied genetics

will play a critical role in d eveloping a pha rma-cology for controlling im mu nologic fun ctions,since it provides the only apparent means of synthesizing man y of the agents that will com-prise immunopharmacology.

ANTIG ENS (VACCINES)

One early dramatic benefit should be in thearea of vaccination, where genetic technologiesmay lead to the production of harmless sub-stances capable of eliciting specific defensesagainst various stubborn infectious diseases.

Vaccination provides effective immunity by

introducing relatively harmless antigens intothe immu ne system thereby allowing the bodyto establish, in advance, adequate levels of anti-body an d a p rimed p opu lation of cells that cangrow when the antigen reappears in its virulentform. Obviously, however, the vaccination itself should not be dangerous. As a result, severalmethods have been d eveloped over the p ast twocenturies to modify the virulence of micro-orga-nisms used in vaccines without d estroying theirability to trigger the production of antibodies.(See Tech. N ote 6, p . 80.)

Novel pure vaccines based on antigens syn-thesized by rDNA have been prop osed to fightcommu nicable d iseases like m alaria, wh ich haveresisted classical preventive efforts. Pure vac-cines have always been scarce; if they wereavailable, they might reduce the adverse effects

of conventional vaccines and change the m eth-ods and the dosages in which vaccines areadministered.

Some vaccines are directed against toxic pro-teins (like the diphtheria toxin produced by

some organisms), prepa ring the body to n eutral-ize them. Molecular cloning might make it pos-sible to produce inactivated toxins, or betternonvirulent fragments of toxins, by means of micro-organisms that are incapable of servingas disease-causing organisms.

Immu nity conferred by live vaccines invari-ably exceeds that conferred by nonliving anti-genic material–possibly because a living micro-organism creates more antigen over a longerperiod of time, providing continuous “boostershots.” Engineered m icro-organisms might be-

come prod uctive sources of high-potency anti-gen, offering far larger, more sustained, dosesof vaccine without the side-effects from the con-taminants found in those vaccines that consistof killed micro-organisms.

How ever, it is clear that formid able Federalregulatory requirements would have to be metbefore permission is granted for a novel livingorganism to be injected into human subjects.Because of problems encountered with live vac-cines, the most likely application will lie in thearea of killed vaccines (often using only parts of

micro-organisms).

It is imp ossible in the scope of this report todiscuss the p ros, cons, and consequences of de-veloping a vaccine for each v iral disease. How -ever, the most commercially important are theinfluenza vaccines, with an average of 20.8 mil-lion doses given per year from 1973 to 1975–asmaller num ber than the 25.0 million d oses peryear of polio vaccine, but more profitable.

Influenza is caused by a virus that has re-mained uncontrolled largely because of the fre-

quency with which it can mutate and change itsantigenic structur es. It has been suggested thatantigenic protein genes for influenza could bekept in a “gene bank” and u sed when n eeded. Inaddition, the genetic code for several antigenscould be introduced into an organism such as E.




coli, so that a vaccine with several antigensmight be prod uced in one fermentation.

1

Two m ore viral diseases deserve at least brief comment. Approximately 800 million doses of foot-and-mouth disease virus (FMDV) vaccineare annually used worldwide, making it the

largest volume vaccine p rodu ced. This vaccinemust be given frequently to livestock in areaswhere the disease is endemic, which includesmost of the world outside of North America.The present methods of producing the vaccinerequire that enormous qu antities of hazardousvirus be contained. Many outbreaks are attrib-uted to incompletely inactivated vaccine or tothe escape of the virus from factories. (Seefigure 23.)

Molecular cloning of the antigen could pro-duce a stable vaccine at considerably less ex-

pense, without the risk of the virus escaping. Onthe basis of that p otential, RAC has approv ed a joint p rogram between th e U.S. Departmen t of Agriculture (USDA) and Genentech to clonepieces of the FMDV genome to prod uce pu re an-tigen. The RAC decision marked the first excep-tion to th e NIH prohibition against cloning DN Athat is derived from a virulent pathogen.

2FMDV.

vaccine made by molecular cloning will prob-ably be distributed commercially by 1985, al-though not in the United States. It will be thefirst vaccine to achieve that status, and illus-trates the potential veterinary uses of genetic

technologies.

Hep atitis has also received significant atten-tion. Vaccines against viral hepatitis, which af-fects some 300,000 Americans each year, maybe produced by molecular cloning. This diseaseis second only to tuberculosis as a cause of death among reportable infectious diseases. R isextremely difficult to cultivate the causativeagents. Hepatitis A has a good chance of beingthe first human viral disease for which the in-itial preparation of experimental vaccine will in-volve molecular cloning. A vaccine against hepa-

titis B, mad e from the blood of chronic carriers,

I For other aspects of vaccine p rodu ction see: Office of Techn ol-ogy Assessment, U.S. Congress, working Papers, The impacts of

Generics, vol. 2. (Springfield, Va.: National Technical InformationService, 1981).

‘Ibid.

Figure 23.-Recombinant DNA Strategy for MakingFoot-and-Mouth Disease Vaccine

I

Growing E. co/i bacteria may produce VP3 for use as vaccinefor foot-and-mouth disease. No virus or infectious RNA is

produced by the harmless bacteria strain.

q VP3 is the protein from the shell of the virus, which can actas a vaccine for immunizing livestock against foot-and-mouth disease. The idea outlined above is to make this VP3protein without making any virus or infectious RNA.


is in the testing stage, but cloning is being in-vestigated as a better source of an appropriateantigen. The causative agent for a third form of hepatitis has not even been iden tified. Since at

least 16 million U.S. citizens are estimated to beat high risk of contracting hepatitis, there iskeen interest in the development of vaccinesamong academic and industrial researchers.

3

31b i d .

76-565 0 - 81 - 6




More hyp othetically, molecular cloning m aylead to three other uses of antigens as well: vac-cination against parasites, such as malaria andhookworm (see Tech. Note 7, p. 80.); immuniza-tion in connection with cancer treatment; andcounteracting abnormal antibodies, which aremade against normal tissues in the so-called“autoimmune diseases, ” such as multiple sclero-sis. (See Tech. N ote 8, p . 81.)

INTERFERON

Interferon are glycoproteins normally madeby a variety of cells in response to viral infec-tion. All int erferon (see Tech. Note 9, p. 81) caninduce an antiviral state in susceptible cells. Inaddition, interferon has been found to have atleast 15 other biochemical effects, most of which involve other elements of the immunesystem.

promising preliminary studies have sup-ported the use of interferon in the treatment of such viral diseases as rabies, hepatitis, varicella-zoster (shingles), and various herpes infections.To date, the effect of interferon has been farmore impressive as a prophylactic than as atherapeutic agent. The interferon produced by

Genentech, for examp le, has been show n to pro-tect squ irrel monkeys from infection by the le-thal m yocarditis virus. Once interferon is avail-able in qu antity, large-scale tests on hu man pop -ulations can be conducted to confirm its ef-ficacy in m an.

Several production techniques are being ex-plored. (See Tech. Note 10, p. 81.) Extraction of interferon from leukocytes (white blood cells),the current m ethod of choice, may have to com-pete with tissue culture production as well asrDN A. (See table 5.)

Recombinant DNA is widely regarded as thekey to mass production of interferon, andimportant initial successes have already beenachieved. Each of the four major biotechnologycompanies is working on improved productionmethod s, and all have reported some success.

An enormou s amoun t remains to be learnedabout the interferon system. It now appearsthat the interferon are simply one of manyfamilies of molecules involved in physiologicalregulation of response to disease. Only nowhave m olecular biology and gen etics made th eirstudy—and perhaps their use—possible.

Table 5.—Summary of Potential Methods for Interferon Production

Types of Potentialinterferon for Present projected Potential for

Means of Production produced scale-up ($/106units) Problems improvement

“Buffy coat” leukocytes leukocyte, 95% No 50 –fibroblast, 5%

Lymphoblastoid cells leukocyte, 80% Yes — =25

fibroblast, 20%

Fibroblasts fibroblast Yes 43-200 = 1-10

Recombinant DNA leukocyte orfibroblast

Yes — z 1-10

—lack of scale up

—pathogen contamination —poor yields —cells derived from tumor

—cell culture —economic competition

with recombinant DNA

—does not produceinterferon

—in vitro drug stability —poor yields

—minimal

—improved yields —expression of

fibroblastinterferon

—improved yields —improved cell-

culturetechnology

—expression ofleukocyte-typeinterferon

—improved yields

—modified

interferon —drug approval —possible economic

competition with fibroblastcell production





The interferon are pr esently receiving atten-tion largely because studies in Swed en and theUnited States stimulated the appropriation of $5.4 million by the American Cancer Society(ACS) for expanded clinical trials in the treat-ment of cancer. That commitment by the non-

profit ACS–the greatest by far in its history–was followed by a boost in NIH fund ing for in-terferon research from $7.7 million to $19.9million for fiscal year 1980. Much of the cost of interferon research is allotted to pr ocuring theglycopep tide. Initially, the ACS bough t 40 billionunits of leukocyte interferon from the FinnishRed Cross for $50 per million units. In March1980, Warner-Lambert was awarded a contractto supply the National Cancer Institute (NCI)with 50 billion units of leukocyte interferonwithin the next 2 years at an average price of $18 per million units. NCI is also planning to

purchase 50 billion units each of fibroblast andlymphoblastoid interferon.

The bulk of the NIH funding is included inNCI’s new Biological Response Modifier (BRM)program—interferon accounts for $13.9 millionof the $34.1 million allocated for BRM work infiscal year 1980. (NCI expenditures on inter-feron in 1979 were $2.6 million, 19 percent of the amount budgeted for 1980.) Other impor-tant elements of that BRM program concernimmunoproteins known as lymphokines andthymic hormones, for which molecular genetics

has m ajor imp lications. The program is aimed atidentifying and testing molecules that controlthe activities of different cell types.

LYMPH OKIN ES A ND CYTOKIN ES

Lymphokines and cytokines are regulatorymolecules that have begun to emerge from theobscure fringes of immunology in the past 10years. (Interferon is generally considered a lym-phokine that has been characterized sufficientlyto deserve ind epend ent status.)

Lymphokines are biologically active solublefactors p rodu ced by w hite blood cells. Stud iedin dep th only w ithin the last 15 years, they arebeing imp licated at virtually every stage in thecomplex series of events that make up the im-mune response. They now include about 100different comp ound s. Cytokines, wh ich have ef-

fects similar to lymphokines, include severalcomp ound s associated w ith the thymu s gland,referred to as thymic hormones.

4

In 1979, the BRM subcommittee concludedthat several of these agents probably have greatpotential for cancer treatment. Nevertheless,adequate quantities for laboratory and clinicaltesting of many of them will probably not beavailable until the p roblems of prod ucing glyco-proteins by m olecular cloning are overcome. Nosystem is currently available for the industrialproduction of glycoproteins, although yeastsmay prove to be the most useful micro-orga-nisms.

ANTIBODIES

Antibodies are the best known and most ex-ploited protein components of the immune sys-tem. Until recently, all antibodies were obtained

from the blood of hu man s or animals; and theywere often impure. Within the past 5 years,however, it has become possible to produce an-tibodies from cells in culture, and to achievelevels of pu rity previously un attainable. As w ithprevious advances in antibody technology, re-searchers are examining ways to put this newlevel of purity to use. There have been hun-dreds, if not thousands, of examples of newdiagnostic and research methods, new methodsof purification, and new therapies publishedwithin the first 3 years that the technique hasbeen available. (See Tech, N ote 11, p. 81.)

This high level of pu rity was attained by thedevelopment of monoclinal antibodies. Theseantibodies that recognize only one kind of anti-gen were the u nanticipated fruit of fund amen-tal immunological research conducted by Drs.Caesar Milstein an d Georges Kohler at the Med-ical Research Cou ncil in England in 1975. Theyfused tw o types of cells–myeloma and plasma-spleen cells—to form hybridomas that producethe monoclinal antibodies. (See Tech. Note 12,p. 81.) Not only are the antibodies specific, butbecause the hybridomas can be grown in mass

culture, a virtually limitless supply is available.

The most imm ediate med ical app lication formonoclinal antibodies lies in diagnostic testing.

4For 40 of the best characterized cytokines, see footnote 1, p.

69.



72 . Impacts of Applied Genetics—Micro-Organ/SmS, Plants, and Animals

Over the past 20 years, large segments of thediagnostic and clinical laboratory industrieshave sprun g up to detect and quantify particu-lar substances in specimens. Because monoclon-al antibod ies are so specific, hybridomas seemcertain to r eplace animals as the sou rce of anti-

bodies for virtually all diagnosis and monitor-ing. Their use will not only improve the accu-racy of tests and decrease development costs,but should result in a more uniform product.

Today, such assays are used to:

determine hormone levels in order toassess the proper functioning of an endo-crine gland or the inappropriate produc-tion of a hormone by a tum or;detect certain proteins, the presence of which has been found to correlate with atum or or w ith a specific prenatal cond ition;

detect the presence of illicit drugs in a per-son’s blood, or mon itor the blood or tissuelevel of a drug to ensure that the dosageachieves a therapeutic level without ex-ceeding the limits that could cause toxic ef-fects; andidentify microbial p athogens.

The extent of the use of antibodies and thebiochemical prop erties that they can identify issuggested by table 6. No one assay constitutes amajor market, and short product lifetime hasbeen characteristic of this business.

Other applications of monoclinal antibodiesinclude:

the imp rovement of the acceptance of kid-ney (and other organ) transplants by injec-tion of the recipient with antibodies againstcertain antigens;passive immunization against an antigen in-volved in rep rodu ction, as a reversible im-mu nological approach to contraception.localizing tumors with tumor-specific anti-bodies (see Tech. Note 13, p. 81); andtargeting cancer cells w ith antibod ies that

have anticancer chemicals attached tothem.

Enzy me s and o the r pro te ins

ENZYMES

Enzymes are involved in virtually every bio-logical process and are w ell-und erstood. Never-theless, despite their potency, versatility, and

diversity, they play a small role in th e pra cticeof medicine today. Therapeutic enzymes ac-counted for American sales of about $70 million

(wholesale) in 1978, but one-half of those salesinvolved the blood-plasma-derived coagulationfactors used to treat hemophilia. Although thefigur e is difficult to estimate, the total nu mberof patients receiving any type of enzyme ther-apy in 1980 probably does not exceed 50,000.

Enzymes cannot be synthesized by conven-tional chemistry. Almost all those presentlyemployed in medicine are extracted fromhuman blood, urine, or organs, or are produced

by m icro-organisms. Already the p ossibility of using rDN A clones as the source of enzym es—primarily to reduce the cost of production—isbeing explored.

How ever, problems associated w ith the use of nonhu man enzymes (such as immun e and feb-rile responses) and the scarcity of human en-zymes, have hindered research, development,and clinical exploitation of enzymes for ther-apeutic purposes. Today, the experimental ge-netic technologies of rDNA and somatic cell fu-sion and culture open the only conceivable

routes to relatively inexpensive production of compatible human enzymes.

The genetic engineering of enzym es is prob-ably the best example of a dilemma that ham-per s the exploitation of rDNA: Withou t a clinicalneed large enough to justify the investment,there is no incentive to produce a product; yetwithout adequate supplies, the therapeutic pos-sibilities cannot be investigated. The substancesthat break this cycle will probably be those thatare already prod uced in quantity from naturaltissue.

The only enzymes administered today aregiven to hemophiliacs–and they are actually




Table 6-Immunoassays .

Analgesics and narcoticsAnileridineAntipyrineCodeineEtorphineFentanyl

MeperidineMethadoneMorphinePentazocine

AntibioticsAmikacinChloramphenicolClindamycinGentamicinIsoniazidPenicillinSisomycinTobramycin

AnticonvulsantsClonazepamPhenytoinPrimidone

Anti-inflammatory agentsColchicineIndomethacinPhenyibutazone

Antineoplastic agentsAdriamycinBleomycinDaunomycinMethotrexate

BronchodilatorsTheophylline

Cardiovascular drugsCardiac glycosides

AcetylstrophanthidinCedilanidDeslanosideDigitoxin

DigoxinGitoxin

Hallucinogenic drugsMescalineTetrahydrocannabi nol

Hypoglycemic agentsButylbiguanideGlibenclamid

InsecticidesAldrinDDTDieldrin

MalathionNarcotic antagonistsCyclazocineNaloxone

Peptide hormonesAngiotensinAnterior pituitaryBradykininGastricHypothalamicIntestinalPancreaticParathyroid

Posterior pituitaryThyroid (calcitonin)

Plant hormoneslndole-3-acetic acidGibberelilic acid

PolyamidesSpermine

ProstaglandinsSedatives and

tranquilizersBarbiturates

BarbitalPentobarbitalPhenobarbital

ChlordiazepoxideChlorpromazineDesmethylimipramine

Diazepam andN-desmethyldiazepam

Methyl digoxinOuabainProscillaridin

DihydroergotaminePropranololQuinidine

CNS stimulantsAmphetamineBenzoyl ecgonine

(cocaine metabolize)MethamphetaminePimozide

DiureticsBumetanide

Hallucinogenic drugsBile acid conjugates

CholylglycineCholyltaurine

CatecholaminesEpinephrineNorepinephrineTyramine

Fibrinopeptides

Fibrinopeptide AFibrinopeptide B

IndolealkylaminesMelatoninSerotonin

Insect hormonesEcdysone

Nucleosides andnucleotides

Cyclic AMPCyclic GMPN

2-Dimethylguanosine

7-MethylguanosinePseudouridineThymidineGlutethimideMethaqualone

Steroid hormonesSkeletal muscle relaxantsd-Tubocurarine

Synthetic peptidesDDAVPSaralasin

Synthetic steroidsAnabolic steroids

Trienbolone acetateAndrogens

FluoxymesteroneEstrogens

DiethylstilbestrolEthinylestradiolMestranol

GlucocorticoidsDexamethasoneMethyl prednisolonePrednisolonePrednisone

MetyraponeProgestins

Medroxyprogesterone

acetateNorethindroneNorethisteroneNorgestrel

ToxinsAflatoxin B,GenisteinNicotine and metabolizesOchratoxin AParalytic shellfish poison

Thyroid hormonesThyroxineTriodothyronine

VitaminsVitamin B12

V i t a m i n D

S O U R C E : ‘“lmmunoassays of Drugs–Comprehensive Immunology,” hrrrnunal Pharmacology, Hadde~ Caffey (cd.) (New York: Plenum Press, 1977) , P. 325.

p r o e n z y m e s , w h i c h a r e c o n v e r t e d t o a c t i v e e n -

z y m e s i n t h e b o d y w h e n n e e d e d . T h e m o s t c o m -

mo n ag en ts a r e ca l led Fac to r VI I I an d Fac to r IX ,

w h i c h a r e f o u n d i n s e r u m a l b u m i n a n d a r e c u r -

r e n t l y e x t r a c t e d f r o m h u m a n b l o o d p l a s m a .

H e m o p h i l i a A a n d H e m o p h i l i a B — a c c o u n t i n g f o r

o v e r 9 0 p e r c e n t o f a l l m a j o r b l e e d i n g d i s o r -

d e r s — a r e c h a r a c t e r i z e d b y a d e f i c i e n c y o f t h e s e

f a c t o r s . S u p p l i e s o f t h e p r o e n z y m e s w i l l e x c e e dd e m a n d w e l l b e y o n d 1 9 8 0 i f t h e h a r v e s t i n g a n d

p r o c e s s i n g o f p l a s m a c o n t i n u e s a s i t h a s . N e v e r -

t h e l e s s , t h e r i s k o f h e p a t i t i s a s s o c i a t e d w i t h t h e

u s e o f h u m a n p l a s m a - d e r i v e d p r o d u c t s i s e x-

t r e m e l y h i g h . O n e r e c e n t s t u d y f o u n d c h r o n i c

h e p a t i t i s i n a s i g n i f i c a n t p e r c e n t a g e o f a s y m p -

t o m a t i c p a t i e n t s t r e a t e d w i t h F a c t o r V I II a n d

Factor IX.

T h e p l a s m a f r a c t i o n a t i o n i n d u s t r y , w h i c h

p r o d u c e s t h e p r o e n z y m e s , i s c u r r e n t l y f a ce d

w i t h e x c e s s c a p a c i t y , i n t e n s e c o m p e t i t i o n , h i g h

p l a s m a c o s t s , a n d t i g h t p r o f i t m a r g i n s .

5

T h e c o s ta n d a v a i l a b i li t y o f a n y o n e p l a s m a p r o t e i n i s

‘ F o r d e t a i l s o f t h e f a c t o r s g o v e r n i n g t h e i n d u s t r y . s e e f o o t n o t e 1 ,

p. 69 .




coupled to the production of the others. Hence,the indu stry wou ld still have to orchestrate theproduction of the other proteins even if just oneof them, such as Factor VIII, becomes a targetfor biological produ ction.

Another enzyme, urokinase, has been tar-

geted for use in removing unw anted blood clots,which lead to strokes, myocardial infarctions,and pu lmonary emboli. Currently, the dru g iseither isolated from urine or produced in tissueculture. (See Tech. Note 14, p. 81.)

Urokinase is thus far the only commercialtherapeutic product derived from m ammaliancell culture. Nevertheless, some calculationssuggest that pr odu ction by E. coli fermentationwou ld have economic advan tages. The costs im-plicit in having to grow cells for 30 days on fetalcalf serum (or its equivalent) or in having to col-

lect and fractionate u rine–as reflected in u roki-nase’s market p rice ($150/ mg at th e manu fac-turer’s level) —should be enough incentive to en-courage research into its p rodu ction. In fact, inApril 1980, Abbott Laboratories disclosed that E. coli had been indu ced to prod uce urokinasethrough plasmid-borne DNA.

The availability of urokinase might be guar-anteed by the new genetic technologies, but itsuse is not. For a variety of reasons, the Amer-ican medical community has not accepted thedrug as readily as have the European and Japa-

nese commu nities. Stud ies to establish the u seof urokinase for deep vein thrombosis, for ex-amp le, are now being cond ucted almost exclu-sively in Eur ope.

6

OTHER PROTEINS

In add ition to the p roteins and polypeptidesalready mentioned, the structural proteins,such as the collagens (the most abundant pro-teins in the body), elastins and keratins (the

compou nd s of extracellular structures like hairand connective tissue), albumins, globulins, anda wide variety of others, may also be susceptibleto genetic engineering. Structural proteins areless likely to be suitable for molecular geneticman ipulations: On the one hand , their size and

eFor additional information about howurokinase came to play arole in th erapy, see footnote 1, p. 69.

complexity exceed the synthetic and analyticcapabilities that will be available in the next fewyears; on the other, either their use in med icinehas yet to be established or material derivedfrom animals appears adequate, as is the casewith collagen, for which uses are emerging.

Plasma, the fluid portion of the blood, con-tains about 10 percent solids, most of which areproteins. During World War II, a simple pro-cedure w as developed to separate the variouscomponents. It is still used today.

Serum albumin is the smallest of the mainplasma proteins but it constitutes about half of plasma’s total mass. Its major therapeutic use isto reverse the effects of shock. p It is a reason-able cand idate for m olecular cloning, althoughits relatively high molecular weight complicatespurification, and its commercial value is rela-

tively low. The market value of normal serumalbumin is approximately $3/ g, but the volumeis such that domestic sales exceed $150 million.Including exports, annual production is in therange of 100,000 kg.

Normal serum albumin for treating shock isalready regarded as too expensive comparedwith alternative treatments, to expand its usewould require a lower price. On the other hand,the Federal Government—and especially the De-partment of Defense—might disregard the im-med iate economic prosp ects and conclude that

having a source of hum an serum albumin thatdoes not depend on p ayments to blood d onorsmight be in the national interest. Since many na-tions imp ort serum albumin, produ cts d erivedfrom m olecular cloning could be exported.

Serum albumin is presently the principalprod uct of blood p lasma fractionation, a changein the w ay it is manu factured wou ld significant-ly affect that industry. Because a number of other products (such as clotting factors) are alsoderived from fractionation, a growth in theneed for plasma-derived album in could h ave asignificant impact on the availability and the

cost of these byproducts.

7For a detailed discussion of the costs and benefits of using albu -

min and the structure of the industry, see footnote 1, p. 69,



Ch. 4—The Pharmaceutical industry . 75

Ant ib io t ics

Antimicrobial agents for the treatment of in-fectious diseases have been the largest sellingprescr ipt ion pharmaceuticals in the world forthe past three decades. Most of these agents are

antibiotics—antimicrobials naturally producedby mic ro-organisms ra the r than by chemica lsynthesis or by isolation from higher organisms.However, one major antibiotic, chlorampheni-col—originally produced by a micro-organism,is now synthesized by chemical methods. Thefield of antibiotics, in fact, provides most of theprecedent for employing microbial fermenta-tion to produce useful medical substances. TheU n i t e d S t a t e s h a s b e e n p r o m i n e n t i n t h e i rdevelopment, production, and marketing, withthe result that American companies account forabout half of the roughly $5 billion worth of an-

t imicrobia l agents sold worldwide each year .The American market share has been growingas new antibiot ics are developed and intro-duced every year.

For 30 ye ar s,high-yielding, antibiotic-pro-ducing micro-organisms have been identified byselection from among mutant strains. Initially,organisms producing new antibiot ics are iso-lated by soil sampling and other broad screen-ing efforts. They are then cultured in the lab-oratory, and efforts are made to improve theirproductivity.

Antibiotics are complex, usually nonprotein,substances, which are general ly the end prod-ucts of a series of biological steps. While knowl-edge of molecular details in metabolism hasmade some difference, not a single antibiotichas had its complete biosynthetic pathway eluci-dated. This is partly because there is no singlegene that can be isolated to produce an antibi-otic. However, mutations can be induced withinthe original micro-organism so that the level of production can be increased.

Other methods can also increase production,

and possibly create new antibiotics. Microbialmating, for example , which leads to naturalrecombination, has been widely investigated asa way of developing vigorous, high-yielding an-tibiotic producers. However, its use has b e e nlimited by the mating incompatibility of many

industr ia l ly important higher fungi , the prence of chromosomal aberrations in micro-ornisms improved by mutation, and a numberothe r problems. Fur the rmore , na tura l r ecbination is most advantageous when strains extremely diverse origins are mated; the pro

prietary secrets protecting commercial strausually prevent the sort of divergent “competitor” strains most likely to produce vigorouhybrids from being brought together.

The technique of protoplasm or cell fusioprovides a convenient method for establishingrecombinant sys tem in s t ra ins , spec ies , agenera that lack an efficient natural meansmating. For example, as many as four strainsthe antibiot ic-producing bacter ium Streptomces have been fused together in a single step tyield recombinant that inherit genes from f

parents. The technique is applicable to neaall antibiotic producers. It will help combine benefits developed in divergent lines by muttion and selection.

In addition, researchers have compared tquality of an antibiotic-producing fungus, Cep alosporimn acremonium, produced by mating toone produced by protoplasm fusion. (See TecNote 15, p. 82.) They concluded that protoplasfusion was far superior for that purpose. Whis more, protoplasm fusion can give rise to hundreds of recombinants–including one isolathat consistently produced the antibiotic cep

alosporin C in 40 percent greater yield than best producer among its parents–without losing that parent strain’s rare capacity to use

organic sulfate, rather than expensive metnine, as a source of sulfur. It also acquired thrapid growth and sporulation characteristicsits less-productive parent. Thus, desirable atributes from different parents were combinin an important industr ia l organism that proved resistant to conventional crossing.

Even more significant are the possibilities fpreparation by protoplasm fusion between di

f e r e n t s p e c i e s o r g e n e r a o f h y b r i d s t r a iwhich could have unique biosynthetic capacties. One group is reported to have isolated anovel antibiotic, clearly not produced by eitheparent , in an organism created through fusof actinomycete protoplasts, (See Tech. Note 1




p. 82.) The value of protoplasm fusion, therefore,lies in p otentially broadening the gene pool.

Protop lasm fusion is genetic recombina tion ona large scale, Instead of one or a few genes be-ing transferred across genus and species bar-riers, entire sets of genes can be moved. Success

is not assured, however; a weakness today is theinherited instability of the “fused” clones. Thepreservation of traits and long-range stabilityhas yet to be resolved. Furthermore, it seemsthat one of the most daunting problems isscreening—determining what to look for andhow to recognize it. (See Tech. Note 17, p. 82.)

Recombinant DNA techniques are also beingexamined for their ability to improve strains,Many potentially useful antibiotics do not reachtheir commercial potential because the micro-organisms cannot be induced to produce suffi-

cient quantities by traditional methods. The syn-thesis of certain antibiotics is controlled byplasmids, and it is believed th at some plasmidsmay nonspecifically enhance antibiotic produc-tion and excretion.

It may also be possible to transfer as a grou p,all the genes needed to produce an antibioticinto a new host. However, increasing the num -ber of copies of critical genes by phage or plas-mid transfer has yet to be achieved in antibiotic-prod ucing organisms because little is know n of the potential vectors. The genetic systems of commercial strains will have to be understoodbefore the newer genetic engineering ap-proaches can be used . Genetic maps have beenpublished for only 3 of the 24 or more indus-trially useful bacteria.

Since 2,000 of the 2,400 know n an tibiotics areproduced by Streptomyces, that is the genus of greatest interest to the pharmaceutical indus-try. Probably every company conducting re-search on Streptomyces is developing vectors,but little of the industrial work has been re-vealed to date.

N o n p r o t ei n p h a r m a c e u t i c a l s

In both sales and quan tity, over 80 percent of the pharmaceuticals produced today are notmade of protein. Instead, they consist of a varie-

ty of organic chemical entities. These drugs, ex-cept for antibiotics, are either extracted fromsome natural plant or animal source or are syn-thesized chemically.

Some of the raw materials for pharmaceuti-cals are also obtained from p lants; micro-orga-nisms are then used to convert the material touseful drugs in one or two enzymatic steps.Such conversions are common for steroid hor-mones.

In 1949, when cortisone was found to be auseful agent in the treatment of arthritis, thedemand for the d rug could not be met since nopractical method for large-scale p roduction ex-isted. The chemical synthesis was complicatedand very expensive. In the early and mid-1950’s,many investigators reported the microbialtransformation of several intermediates to com-

poun ds that correspond ed to the chemical syn-thetic scheme. By saving many chemical stepsand achieving higher yields, manufacturersmanaged to reduce the price of steroids to alevel where they were a marketable commodi-ty. A conversion of progesterone, for example,dropped the price of cortisone from $200 to$6/g in 1949. Through f u r the r im pr ove m e n t s ,the price dropped to less than $1/ g. The 1980pr ice is $0.46/ g.

Developm ents based on genetic techniques toincrease the pr odu ction and secretion of key en-

zymes could su bstantially improve the econom-ics of some presently inefficient processes. Cur-rently, assessments are being carried out byvarious companies to determine which of themany nonprotein pharmaceuticals can be man-ufactured more readily or more economicallyby biological means.

Approximately 90 percent of the pharmaceu-ticals used in the treatment of hypertension areobtained from plants, as well as are miscel-laneous cardiovascular drugs. Morphine andimportant vasodilators are obtained from the

opium poppy, Papaver somniferum. All thesechemical substances are produced by a series of enzymes that are coded by corresponding genesin the whole plant. The long-term possibility(over 10 years) of using fermentation m ethodswill dep end on identifying the important gen es.




The genes that are transferred from plant to

bacteria must obviously be determined on a

case-by-case basis. The case study on acetamino-

phen (the active ingredient in analgesics such as

Tylenol) demonstrates the steps in such a feasi-

bility study. (See app. I-A.)

The first step in such a stud y is to determinewh ether and w here enzymes exist to carry outthe necessary transformation for a given prod-uct. Acetaminophen for instance, can be m adefrom aniline, a relatively inexpensive startingmaterial. The two necessary enzymes can be

found in several fun gi. Either the enzym es canbe isolated and used directly in a two-step con-version or the genes for both enzymes can betransferred into an organism th at can carry outthe entire conversion by itself.

Given the cost assumptions outlined in thecase stud y and the assum ptions on th e efficien-cy of converting aniline to acetaminophen , thecost of producing the drug by fermentationcould be 20 percent lower than production bychemical synthesis.

I m p a c t s

Genetic technologies can help provide a varie-ty of pharmaceutical products, many of which

have been identified in this report. But the tech-nologies cannot gu arantee how a p rodu ct w illbe used or even whether it will be used at all.The pharmaceuticals discussed have illustratedthe kinds of major economic, technical, social,and legal constraints that will play a role in theapplication of genetic technologies.

Clearly, the major direct impacts of genetictechnologies will be felt primarily throu gh th etype of products they bring to market. Never-theless, each new ph armaceutical will offer itsown spectrum and magnitude of impacts. Tech-

nically, genetic engineering may lead to the pro-duction of growth hormone and interferon withequal likelihood; but if the pa tient p opu lation isa thou sand fold h igher for interferon, and if itstherap eutic effect is to alleviate pain and lowerthe cancer mortality rate, its impact w ill be sig-nificantly greater.

Many hormones and h um an proteins cannotbe extensively studied because they are stilleither unavailable or too expensive. Until thephysiological properties of a hormone areunderstood, its therapeutic values remain u n -

kn own. Recombinan t DNA techniq ues are beingused to overcome this circular problem. In onelaboratory, somatostatin is being used as a re-

search tool to study the regulation of the hor-

monal milieu of burn patients. A single experi-

ment may use as much as 25 mg of the hor-

mone, which, as a produ ct of solid state chem-ical synthesis, costs as much as $l2,000. Re-

du cing its cost wou ld allow for more extensiveresearch on its physiological and therapeuticqualities.

By m aking a ph armaceutical available, genet-ic engineering can have two types of impacts.First, pharmaceuticals that already have med-ical promise will be available for testing. For ex-ample, interferon can be tested for its efficacyin cancer and viral therapy, and human growthhorm one can be evaluated for its ability to healwounds. For these medical conditions, the in-direct, societal impact of applied genetics could

be widespread.Second , other pharmacologically active su b-

stances that have no present use will be avail-able in sufficient quantities and at a low enoughcost to enable researchers to explore their possi-bilities, thus creating the potential for totallynew therapies. Genetic technologies can makeavailable for example, cell regulatory proteins, aclass of molecules that control gene activity andthat is found in only minute quantities in thebody. The cytokines and lymphokines typify thecountless rare molecules involved in regu lation,communication, and defense of the body tomaintain health. Now, for the first time, genetictechnologies make it possible to recognize, iso-late, characterize, and prod uce these p roteins.

The potential importance of this class of phar-maceuticals—the new cell regulatory mole-




cules—is underscored by the fact that half of the 22 active INDs for new molecular entitiesthat have been rated by FDA as promising im-portant therapeutic gains are in the Metabolicand Endocrine Division, which oversees suchdrugs. It is reasonable to anticipate that they

will be employed to treat cancer, to prevent orcombat infections, to facilitate transplantationof organs and skin, and to treat allergies andother diseases in which the immune system hasturned against the organism to w hich it belongs.(See table 7.)

At the very least, even if immed iate medicaluses cannot be found for any of these com-pounds, their indirect impact on medical re-search is assu red. For the first time, almost anybiological phenom enon of medical interest canbe explored at the cellular level by the appli-

cable 7.–Diseases Amenable to Drugs Produced byGenetic Engineering in the Pharmaceutical Industry

Drug potentially produced byDisease or condition genetically engineered organism

DiabeteseAtherosclerosis

Virus diseasesInfluenzaHepatitisPolioHerpesCommon cold

Cancer

AnovulationDwarfism a

PainWounds and burnsInflammation,

rheumatic diseasesa

Bone disorders, e.g.,Paget’s disease

a

Nerve damageAnemia, hemorrhage

HemophiliaBlood clotsa

Shocka

Immune disorders

InsulinPlatelet-derived growth factor(PDGF)

Interferon

InterferonHodgkin’s diseaseLeukemiaBreast cancer

Human chorionic gonadotropinHuman growth hormoneEnkephalins and endorphinsHuman growth hormone

Adrenocorticotrophic hormone(ACTH)

Calcitonin and parathyroidhormone

Nerve growth factor (NGF)Erythropoietin

Factor Vlll and Factor IXUrokinaseSerum albuminCytokines

alndicates dlsaaaes currently treated by the drugs listed.


cation of available scientific tools. These newmolecules are valuable tools for dissecting thestructure and function of the cell. The knowl-edge gained may lead to the development of new therapies or preventive measures fordiseases.

The increased availability of new vaccinesmight also have serious consequences. But theextent to w hich molecular cloning will provideuseful vaccines for intractable diseases is stillun know n. For some widespread diseases, suchas amebic dysentery, not enough is knownabout the interaction between the micro-orga-nism and the patient to help researchers designa rational plan of attack. For others, such astrachoma, malaria, hepatitis, and influenza,there is only preliminary experimental evidencethat a useful vaccine could be produced. (Seetable 8.) To date, the vaccine that is most likely

to have an immediate impact combats foot-and-mou th d isease in veterinary m edicine. There islittle dou bt however, that shou ld any one of thevaccines for hu man diseases become available,the societal, economic, and political conse-quences of a decrease in morbidity and mortali-ty w ould be significant. Many of these diseasesare particularly prevalent in less-developedcountries. The effects of developing vaccines

Table 8.-Major Diseases for Which VaccinesNeed To Be Developed

Parasitic diseasesHookwormTrachomaMalariaSchistosomiasis (river blindness)Sleeping sickness

virusesHepatitisInfluenzaFoot-and-mouth disease (for cloven-hoofed animals)Newcastle disease virus (for poultry)Herpes simplexMumpsMeaslesCommon cold rhinovirusesVaricella-zoster (shingles)

8ecterJaDysenteryTyphoid feverCholeraTraveller’s diarrhea





Clearly, there is no simple formula to identify Given the assumptions described, the immedi-all the imp acts of applied genetics on the phar - ate direct economic impact of using genetic ma-maceutical industry. Even projections of eco- nipulation in the industry, measured as sales,

nomic impacts must remain crude estimates. can be estimated in the billions of dollars, with

Nevertheless, the degree to which genetic engi- the indirect impacts (sales for suppliers, savings

neering and fermentation technologies might due to decreased sick

potentially account for drug prod uction in spe- several times th at value.cific categories is projected in appendix I-B.

days, etc.) reaching

T e ch n i ca l n o t e s

1. Many h ormones are simply chains of am ino acids (poly -peptides); some are polypeptides that have been mod-ified by th e attachm ent of carbohyd rates (glycopep -tides). Hormon es usually trigger events in cells remotefrom the cells that produced them. Some act overrelatively short distances —between segments in thebrain, or in glands closely linked to the brain, othersact on d istant sites in tissues throughout the body.

2. For peptides ab out 30 AA in length, the cost may ap-

3.

4.

preach $1 per mg as the volume approaches the kilo-gram level–a level of deman d rarely existing today b utlikely to be generated by work in progress. Today, thecost of the 32 AA polypeptide, calcitonin, which is syn-thesized chemically and m arketed as a pharmaceuticalprodu ct by Armour, is probably in the range of $20 permg, since the wh olesale price in vials containing ap -proxim ately 0.15 mg is ab out $85/mg. (That p rice is anedu cated gu ess, since such costs are closely guardedsecrets and since the price of a pharmaceutical in-cludes so many variables that the cost of the agentitself is a small consideration. )In add ition to those h elped by the National PituitaryAgen cy, ano ther 100 to 400 patients ar e treated w ith

hGH from comercial sources. The commercial priceis approximately $15 per unit (roughly $30/mg). Theprod uction cost at the N ational Pituitary Agency isabout $0.75/unit ($1.50/mg). The National PituitaryAgency produces 650,000 international units (lU)

(about 325 g) of hG H, along with the thyroid-stimulat-ing horm one, prolactin, and other hormon es, fromabout 60,000 human pituitaries collected each year.That is enough hG H both for the current demand andfor perhaps another 100 hypopituitary patients.Workers at the How ard Hu ghes Medical Institute of the University of California, San Francisco, isolatedmessenger RNA from a hu man p ituitary tumor andconverted it into a DN A-sequence that could be pu t intoE. coli. The sequence, however, was a mixture of hGH

and non-hGH material. It has been reported that Eli Lil-ly & Co., which has provided some grant m oney to theInstitute, has obtained a license to the patents relatingto this work. Grants from the N ational Institutes of Health and the National Science Found ation were alsoacknowledged in the pu blication.

At practically the same tim e, researchers at Gen en-tech, in conjunction with their associates at City of Hope N ational Medical Center disclosed the p roductionof an hGH analog. This was the first time that a humanpolypep tide was directly expressed in E. coli in func-tional form. The work was supported by Kabi Gen AB,and Kabi has the m arketing rights.

The level of hGH production reported in the scientif-ic account of the Genentech work was on the sameorder as that reported for the insulin fragments—approximately 186,000 hGH molecules per cell—a levelthat might be competitive even before efforts are madeto increase yield. Genen tech stresses the poin t that de-sign, rather than classical mutation and selection, is thelogical way to improve the system, since the hormone’s“bluep rint” is incorporated in a plasmid th at can bemoved between strains of E. coli, between sp ecies, oreven from sim ple b acteria into more complex orga-nisms, such as yeast.

5. Since erythropoietin is a glycoprotein, it may not befeasible to synthesize the active hormone with present-ly available rDN A techniques.

6. Antigens are surface componen ts of pathogenic orga-

nisms, toxins, or other proteins secreted by path ogenicmicro-organisms. They are also the specific counter-parts of antibodies: antibodies are formed by thebody’s immune system in response to their presence.Antibodies are synthesized b y white b lood cells and arecreated in such a way that they are uniquely struc-tured to bind to specific antigens.

7. Many of the most devastating infectious diseases in-volve comp lex parasites that refuse to grow u nd er lab-oratory conditions. The first cultivation of th e mostmalignant of th e species of protozoa that causes ma-laria, using h uman red blood cells, was described in1976 by a Rockefeller University p arasitologist, WilliamRager. Experimental immunogens were prepared andshowed p romise in monkeys, but concern about the ex-

istence of the red blood cell remnan ts—wh ich couldgive rise to autoimm un e reactions–curtailed the p ros-pect for mak ing practical vaccines by that rou te. Sever-al biotechnology firms are currently trying to synthe-size malaria antigens b y molecular cloning. This effortmay p roduce technical solutions to such scourges as



Ch. 4—The Pharmaceutical industry q 81

schistosomiasis (bilharzia), filariasis (onchocerciasisand elephantiasis), Ieshmaniasis, hookworm, amebic in-fections, and trypanosomiasis (sleeping sickness andChagas’ disease).

8. Another p otential use of antigens is suggested by th eexperimental treatmen t of stage I lung cancer patientswith vaccines prepared from pu rified hu man lun g

cancer antigens, which app ears to substantially pro-long survival. And the Salk Institute is expanding clin-ical trials in which a procine myelin p rotein preparedby Eli Lilly & Co. is injected into mu ltiple sclerosis pa-tients to mop up the antimyelin antibodies that thosepatien ts are producing. Fifteen to forty-two g of myelinhave been in jected without adverse effects, suggestinga new therapeutic approach to auto-immune diseases.The protein appears to suppress the symptoms of ex-perimental allergic encephalomyelitis, an animal dis-ease resembling multiple sclerosis. Should this re-search succeed, the u se of molecular clones to produ cehuman protein antigens seems inevitable.

9. There are at least two distinct kinds of “classical” inter-ferons—leukocyte interferon and fibroblast interferon,so-called for the types of cells from which th ey are ob-tained. A third kin d, called lymphoblastoid because it isprodu ced from cells derived from a Burk itt’s lymph o-ma, appears to be a mixture of the other two inter-feron. All produce the antiviral state and are inducedby viruses. A fourth kind, kn own as “imm une” inter-feron, is produced by lymp hocytes. Some evidence in-dicates that it may be a more potent antitumor agent

t h a n t h e c l a s s i c a l t y p e s . C u r r e n t l y , i n t e r f e r o n i s o b -

tained chiefly from white blood cells (leukocytes) fromthe blood bank in Helsinki that serves all of Finland, orfrom fibroblasts grown in cell culture.

10. Recently, G. D. Searle & Co. annou nced th at new tech-nology developed at its R&D facility in England has in-creased the yield of fibroblast interferon by a factor of 60. On th e basis of this process, Searle expects to sup -

ply material for the first large-scale clinical trial of fibroblast interferon. Abbott Laboratories also recentlyannounced plans to resume production of limitedqu antities of fibroblast interferon for clinical studies itplans to sponsor.

Unlike leuk ocytes and specially treated fibroblasts,wh ich can be used only once, lymph oblasts derivedfrom the tumor Burkitt’s lymphoma grow freely insuspension and produce the least costly interferonpresently obtainable. However, they also produce a dis-advantageous mixture of both leukocyte and fibroblastinterferon. The Burroughs-Wellcome Co. produceslymphoblastoid interferon in 1,000-1 fermenters andhas begu n clinical trials in England , but the U.S. FDAhas generally resisted efforts to make use of p roducts

derived from malignan t cells. It is used extensively inresearch, and FDA is considerin g evidence from Bur-roughs-Wellcome th at may lead to a relaxation of theprohibition, under pressure from the National CancerInstitute.

11. What may be a land mark p atent has been issu ed toHilary Koprowski and Carlos Croce of the Wistar Insti-

tute (for work done under the then Department of Health, Education, and Welfare funding) on the pro-du ction of mon oclinal antibod ies against tumor cells.In a nu mb er of examples, these researchers dem on-strated that an animal can be immu nized with tumorcells, and th at hybridom as derived from that animalwill produ ce antibodies that demon strate a specificity

for the tumor.The final senten ce of the patent text provides the ra-tionale for the use of antibodies in b oth cancer and in-fectious d isease therapies: “If the (tumor) antigen ispresent, the patient can be given an injection of an anti-body as an aid to react with the antigen. ” (U.S.4,172,124.)

12. Myeloma cells grow vigorously in culture and have theun ique characteristic of produ cing large quantities of antibodies. Each spleen cell of the immune type, on theother hand, produces an antibody that recognizes asingle antigen, but these do not grow well in culture.When normal immune spleen cells are fused with mye-loma cells, the resultin g mixture of genetic capacitiesforms a cell, called a “hybridom a,” which displays thedesired characteristics of the parent cells: 1) it secretesthe antibod y specified by the genes of the spleen cell;and 2) it displays the vigorous growth, production, andlongevity that is typical of the m yeloma cell.

13. The use of high-correlation an tibody assays in cancerstudies has only just begun. Antibodies that have beentreated so they can be seen w ith X-rays and that arespecific for a tum or, can b e used early to detect the oc-currence or spread of tumor cells in the bod y. Because

some 785,000 new cancer cases will be detected in

1980 with current diagnostic methods, because cancer

will cause 405,000 deaths, and because early detectionis the major key to improving survival, the implicationsare indeed enormous.

14. In the late 1950’s, Lederle Laboratories marketed apreparation of 95-percent p ure streptok inase (a bac-

terially produced enzyme that dissolves blood clots) forintravenous adm inistration. They withdrew the p rod-uct from the m arket aroun d 1960 because it causedallergic reactions, which dampened clinical enthusiasmfor its therapeutic potential.

The presence in hu man u rine of urokinase, an en-zyme also capable of removing blood clots, was also dis-covered in the early 1950’s. Urokinase was purified,crystallized, and brought into clinical use in the mid-1960’s. From the beginning it was apparent that “an in-tense thrombolytic state could be achieved with amu ch milder coagulation d efect than occurred withstreptokinase; no p yrogenic or allergic reactions w erenoted, and n o antibodies resulted from its administra-tion . . . There did n ot appear to be as great variation in

patient responsiveness.” In 1967-68 and 1970-73, theNational Heart and Lung Institute organized clinicaltrials that compared urokin ase with streptokinase andheparin, an anticoagulant, in the treatment of pul-monary emb olism. The trials indicated that strep-tokinase and urokinase were equivalent and sup eriorto heparin over the short term, although their long-



82 q Impacts of Applied Genetics —Micro-Organisms, Plants, and Animals

term ben efits were not established . Since then, clinicalinvestigation of urokinase has been hampered bydomestic regulatory problems, which have raised thecost of produ ction and restricted its availability in theUnited States.

In January 1978, Abbott Laboratories obtained anew drug application for urokinase and introduced the

produ ct Abbokinase; by that time, however, the salesof urokinase in Japan were already pushing $90 millionper year. Recently, Sterling Dru g has begun mark etinga urokinase product (Breokinase) manufactured byGreen Cross of Japan: “According to Japan ese reports,urokin ase is the first Japan ese-made drug formu lationto receive produ ction an d sales approval from FDA.Green Cross estimates that within 3 years of the startof Sterling’s marketing activities, the value of uro-kin ase exports w ill reach Yen 500 million ($2.12 mil-lion) per month , and considers that its profits from ex-porting a finished product will probably be better thanthose from bu lk dru g sales or the licensing of technol-ogy. ” The Green Cross product is made from humanurine collected through out Korea and Japan, and takesadvantage of technology licensed from Sterling. Ab-

bott’s product, on the other hand, is derived fromkidney-cell culture.

15. Intergeneric hybrids have extremely interesting pos-sibilities. For examp le, it would b e ben eficial to cepha-Iosporin-process technology to combine in one orga-nism the acyltransferase from Penicillium chrysogenum

and the enzymes of C. acremonium, which does not in-corporate side chain precursors onto cephalosporinl ike P. chrysogenum does for pen icillins.

16. Another example of recombin ation betw een sp ecies isthat reported for two sp ecies of fungi, Aspergillus nidu-lans and A. rugulosus, subsequent to protoplasm fusion.

The only report of a successful cross betweengenera using protoplasm fusion technology has been b e-tween the yeasts Candida tropicalis an d Saccario-

mycopsis fibuligera, wh ich took p lace at low frequ encyand gave rise to types intermediate between theparents.

17. An example of screening is provided by the newB-lactam (penicillin-like) antibiotics. Using olderscreening methods, no new B-lactams were foundfrom 1956 until 1972 when a new method was devised.A new series of these antibiotics was thus found.Within the past year, 6 new B-lactams have beencommercialized and at least 12 more are in clinical

trials aroun d th e world. The sales forecasts for thesenew agen ts are estimated to be over $1 billion.



Ch a p t e r 5

Page

Backgrou nd . . . . . . . . . . . . . . . . . . . . . . . . . 85Overview of the Industry . . . . . . . . . . . . . . . . . . 85

Fermentation and the Chemical Industry. . . . . 87New Process In trod uction. . . . . . . . . . . . , , . . . . 89

Characteristics of Biological Production

Technologies. . . . . . . . . . . . . . . . . . . . . . . . . 90Renewable Resources. . . . . . . . . . . . . . . . . . 90Physically M ild er Con di tion s. . . . . . . . . . . . . 91On e-Step Prod uction Meth ods. . . . . . . . . . . . 91Reduced Pollution... . . . . . . . . . . . . . . . . . 91

Indu strial Chemicals That May Be Produced byBiological Technologies. . . . . . . . . . . . . . . . . 92

Fertilizers, Polymers, an d Pe sticid es. . . . . . . . . 94Constraints on Biological Production Techniques 96An Overview of Impacts . . . . . . . . . . . . . . . . . . . 97

Impacts on Other Industries. . . . . . . . . . . . . . . 97Impacts on University Research. . . . . . . . . . . . 98The Social Impacts of Local Industrial Activity . 99Imp acts on Manp ow er. . . . . . . , . . . . . . . . . . . 99

Tables

Table No. Page

9. Data for Commercially Produced AminoAcids. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 88

10.

11.

12.

13.

14.

15.

16.

17.18.19.

Summary of Recent Estimates of PrimaryU.S. Cost Factors in the Prod uction of Lysine Monohydrochloride by Fermentationand Chemical Synthesis. . . . . . . . . . . .Some Commercial Enzymes and Their U

Expansion of Fermentation Into theChemical In dus try, . . . . . . . . . . . . . . .The Potential of Some Major PolymericMaterials for Production Using Biotechnology 95Some Private Companies WithBiotechn ology Program s ., . . . . . . . . . . . . , . 98Distribution of Applied Genetics Activityin Industry . . . . . . . . . . . . . . . , . . . . . . . . . . 100Manpower D istribution of a Firm WithApplied Genetics Activity. . . . . . . . . . . . . . . . 100Index to Fermentation Companies. . . . . . . . . 100Fermentation Products and Producers . . . . . 101U.S. Fermen tation Com pan ies . . . . . . . . . . . . 103

Figures

Figure No. Page

24. Flow of industrial Organic Chemicals FromRaw M aterials to Consu mp tion . . . . . . . . . . . 86

25. Diagram of Alternative Routes to OrganicChemicals , . . . . . . . . . . . . . . . . . . . . . . . . . . 90




Figure 24.-Flow of Industrial Organic Chemicals From Raw Materials to Consumption

Organic resources

80% raw material from petroleum/natural gas

20°/0 raw material from coal, coke, andrenewable resources

SOURCE: U.S. Industrh?l Outlook (W.shlngton, D. C.: Department of Commerce, 1978); Kline Gu/de to Chemioel Industry, Fairfield,N.J., adapted from Tong, 1979.



Ch. 5—The Chemical industry q 87

alone serves as the basic chemical for the manu-facture of half of the largest volum e indu strialchemicals. Each of the steps in a chemical con-version process is controlled by a separate reac-tion, which is often performed by a separatecompany.

Evaluating the competitiveness both of aprocess and of the market is critical for thechemical industry, which is intensive for cap-ital, energy, and raw materials. Its plants uselarge amounts of energy and can cost hundredsof millions of dollars to build, and raw materialcosts are generally 50 to 80 percent of a p rod -uct’s cost. If a biological process can use thesame raw materials and reduce the process costby even 20 percent, or allow the use of inexpen-sive raw ma terials, it could provide th e industrywith a major price break.

F e r m e n t a t i o n a n d

t h e c h em i ca l i n d u s t r y

The production of industrial chemicals byfermentation is not new. Scores of chemicalshave been produced by micro-organisms in thepast, only to be replaced by chemical produc-tion based on p etroleum. In 1946, for examp le,27 percent of the ethyl alcohol in the UnitedStates was produced from grain and grain prod-ucts, 27 percent from molasses, a few percenteach from such materials such as potatoes, pine-

app le juice, cellulose pu lp, and wh ey, and on ly 36 percent from petroleum. Ten years lateralmost 60 percent w as derived from p etroleum.

Even more dramatically, fumaric acid was atone time produced on a commercial scalethrough fermentation, but its biological produ c-tion was stopped when a more economical syn-thesis from benzene w as developed. Frequently,after a fermentation product was discovered,alternative chemical synthetic methods weresoon developed that used inexpensive petro-leum as the raw m aterial.

Nevertheless, for the few chemical entitiesstill prod uced by fermentation, app lied genetics

has contributed to the economic viability of theprocess. The production of citric and lacticacids and various amino acids are among theprocesses that have benefited from genetics.Lactic acid is prod uced both syn thetically andby fermentation. Over the past 10 to 20 years,

manufacture by fermentation has experiencedcompetition from chemical processes.

The organisms used for the prod uction of lac-tic acid are v arious sp ecies of the bacterium Lac- tobacillus. Starting materials may be glucose, su-crose, or lactose (whey). The fermentation perse is efficient, resulting in 90 percent yields, de-pending on the original carbohydrate. Sincemost of the problems in the m anu facture of lac-tic acid lie in the recovery pr ocedu re and not infermentation, few attempts have been made toimprove the industrial processes throughgenetics.

Citric acid is the most important acidulant,and historically has held over 55 to 65 percentof the acidulant market for foods. * It is alsoused in p harm aceuticals and miscellaneous in-du strial applications. It is pr odu ced commercial-ly by the mold Aspergillus niger. Surpr isingly lit-tle work has been p ublished on imp roving citricacid-pr odu cing strains of this m icro-organism.Weight yields of 110 percent have recently beenreported in A. niger mutants obtained by ir-radiating a strain for wh ich a m aximu m yield of 29 percent had been reported.

Amino acids are the building blocks of pro-teins. Twenty of them are incorporated intoproteins manufactured in cells, others servespecialized structural roles, are important meta-bolic intermediates, or are hormones and neu-rotransm itters. All of the am ino acids ar e usedin research and in nutritional preparations,with most being used in the preparation of pharmaceuticals. Three are used in large qu an-tities for two p urp oses: glutam ic acid to m anu -factur e monosod ium glu tamate, wh ich is a fla-

q The other two important acidukmts , or acidifying agents, ar ephosphoric acid (20 to 25 percent) an d maltc acid (S percent),




vor enh ancer particularly in oriental cooking;*and lysine and methionine as animal feed ad-ditives.

Conventional technology for producing glu-tamic acid is based on p ioneering w ork that w assubsequently applied to other amino acids. Theproduction employed microbial strains to pro-du ce amino acids that are not within th eir nor-mal biosynthetic capabilities. This was accom-

“Monosodium glutamate is the sodium salt of glutamic acid. In1978, about 18,000 tonnes were manufactured in the UnitedStates and abou t 11,000 tonnes imported. The food industry con-sumed 97 percent. The fermentation plant of the Stauffer Chem-ical Co. in San Jose, Calif., is the sole U.S. prod ucer. The microbesused in glu tamic acid fermentation (Corynebacferium glutamicum,

C. Iileum, an d Brevibacterium j?avum) produce it in 60 percent of theoretical yield. Thus, there is some b ut n ot great potential forthe use of app lied genetics to improve the yield. Many of the ge-netic approaches have already been thoroughly investigated by in -dustrial scientists.

plished by u sing two methods: I) manipulatingmicrobial growth conditions, and 2) isolatingnat u rally occurring m utants.

Although microbial production of all theamino acids has been stud ied, glutam ic acid an dL-lysine* * are the ones prod uced in significantquantities by fermentation processes. (See table9.) The produ ction of L-lysine is an excellent ex-

“ *The lack of a single amino acid can retard protein synth esis,and th erefore growth, in a mamm al. The limiting amino acid is afunction of the animal an d its feed. The major source of animalfeed in th e United States is soybean meal. The limiting amino acidfor feeding swine is lysine, the limiting amino acid for feedingpoultry is meth ion ine . Because of increased poultry dem and,world demand for Iysine is climbin g, Eurolysine is spend ing $27million to dou ble its produ ction capacity in Amiens , France, to 10thousand ton nes. The Asian and Mid east markets ar e ;st imated toincrease to 3 thousand tonnes in 1985. Some ba r .eria producelysine at over 90 percent of theoretical yield, L:[tle genetic im-provement is likely in this conversion yield, however, significantimprovement can be made in the rate and final concentration.

Table 9.—Data for Commercially Produced Amino Acidsa

Price March Potential for application of1980 (per kg Production 1978 biotechnology (de novo synthesis or

Amino acid pure L) Present source (tonnes) bioconversion; organisms and enzymes)

Alanine. ... .. ... ... ... ... .$ 80

Arginine . . . . . . . . . . . . . . . . . . .Asparagine. . . . . . . . . . . . . . . . .Aspartic acid . . . . . . . . . . . . . . .

Citrulline. . . . . . . . . . . . . . . . . . .Cysteine. . . . . . . . . . . . . . . . . . .Cystine. . . . . . . . . . . . . . . . . . . .DOPA (dihydrophenylalanine) .

Glutamic. . . . . . . . . . . . . . . . . . .Glutamine. . . . . . . . . . . . . . . . . .Histidine. . . . . . . . . . . . . . . . . . .Hydroxyproline . . . . . . . . . . . . .Isoleucine. . . . . . . . . . . . . . . . . .Leucine. . . . . . . . . . . . . . . . . . . .Lysine. . . . . . . . . . . . . . . . . . . . .

Methionine. . . . . . . . . . . . . . . . .

Ornithine . . . . . . . . . . . . . . . . . .Phenylalanine . . . . . . . . . . . . . .

Proline . . . . . . . . . . . . . . . . . . . .Serine. . . . . . . . . . . . . . . . . . . . .Threonine. . . . . . . . . . . . . . . . . .

Tryptophan. . . . . . . . . . . . . . . . .Tyrosine. . . . . . . . . . . . . . . . . . .Valine . . . . . . . . . . . . . . . . . . . . .

285012

2505060

750

455

160280350

55350

265

6055

125320150

1101360

Hydrolysis of protein; 10-50(J) b –chemical synthesis

Gelatin hydrolysis 200-300 (J) Fermentation in JapanExtraction 10- 50(J) –Bioconversion offumaric acid 500-1,000 (J) Bioconversion

10-90 (J) Fermentation in JapanExtract ion 100- 200(J) –Extraction 100- 200(J) –Chemical 100- 200(J) —

Fermentation 10,000-100,000 (J) De novo: Micrococcus gutamicus Extraction 200-300 (J) Fermentation in Japan — 100-200 Fermentation in JapanExtraction from collagen 10-50 —

Extraction —

Fermentation (800A)Chemical (20%)

Chemical from acrolein

—

Chemical frombenzaldehyde

Hydrolysis of gelatin — —

Chemical from indoleExtraction —

aproduction data largely from Japan because of relative Small U.S. production.L

10-50 (J)50-100(J) Fermentation in Japan10,000 (J) (80% by fermentation) De novo:

Corynebacterium glutamicum andBrevibacterium flavum

17,000(D,L) c — 20,000 (D,L) (J)10-50 (J) Fermentation in Japan50-100 (J) Fermentation in Japan

10-50 (J) Fermentation in Japan10-50 (J) Bioconversion in Japan50-10 (J) Fermentation in Japan

55 (J)50- 100(J) -50-100 (J) Fermentation in Japan

‘Japan.C

Dand L forms.

SOURCE: Massachusetts Institute of Technology.



Ch. 5—The Chemical Industry . 89

amp le of the competition betw een chemical and mostly from Japan and South Korea. Recentbiotechnological methods. Fermentation has estimates of primary U.S. cost factors in thebeen gradually replacing its production by competing production methods are summarizedchemical synthesis; in 1980, 80 percent of its in table 10. Fermentation costs are lower for allworldw ide production is expected to be by mi- three components of d irect operating costs:crobes. It is not p rodu ced in the Un ited States, labor, material, and utilities.

which imported about 7,000 tonnes in 1979,

Table 10.—Summary of Recent Estimates of Primary U.S. Cost Factors in the Production ofL= Lysine Monohydrochloride by Fermentation and Chemical Synthesis

Cost factors in Production of 98% L-lysine monohvdrochloride

By fermentation By chemical synthesis

Requirement Estimated 1976 cost Requirement Estimated 1976 cost

(units per unit per unit product (units per unit per unit product

(product) Cents/lb Cents/kg product) Cents/lb Cents/lb

Total Iaborc. . . . . . . . . . . . . . . . . — 8 18 — 9 20

MaterialsMolasses . . . . . . . . . . . . . . . . 44 7 16 —Soy beanmeal, hydrolyzed. . .

— —

0.462 4 9 —

Cyclohexanol. . . . . . . . . . . . . —

— —

— 0.595 17 37Anhydrous ammonia. . . . . . . — — — 0.645 6 14Other chemicals

d. . . . . . . . . . — 7 15 — 4 10

Nutrients and solvents. . . . . — — — — 4 8Packaging, operating, and

mainten ance materials. . . 10 22 — 9 21

Total materials. . . . . . . . . . — 28 62 — 45 90Total utilities . . . . . . . . . . . . . . — 6 12 — 7 16

Total direct operating cost — 42 92 — 56 126Plant overhead, taxes,

and insurance . . . . . . . . — 10 21 — 10 21

Total cash cost . . . . . . . . . — 52 11 — 66 147Depreciation. . . . . . . . . . . — 16 35 — 13 28Interest on working capital — 1 3 — 1 3

Total costg. . . . . . . . . . . — 69 151 — 80 178

aAggumes a 23-percent yield on molasses.bAssumes a 65-percent yield on cyclohexanol.clncludes operating, maintenance, and control laboratory labor.dFor both the proce~~ of fermentation and chemical synthesis, assumed use of hydroch lo ric acid (~ percent) and ammonia (n percent). For fermentation includes dSO

potassium diphosphate, urea, ammonium suifate, calcium carbonate, and magnesium sulfate. For chemical synthesis also includes nitrosyl chloride, sulfuric acid,and a credit for ammonium sulfate byproduct.~otal utiiities for both processes include cooling water, steam process water, and electricity. For chemical synthesia, natural gas is also included.fTen percent ~r year of fixed capital costs for a new 20 million lb per yea r U.S. plant built in 1975 at assumed capital cost of $36.6X 10’ for fOrmOntatiOn and $32.5x 10’

for chemical synthesis exclusive of land costs.

SOURCE: Stanford Research Institute, Chemical Economics Handbook 563:3401, May 1979.

N e w p r o c e s s i n t r o d u c t i o n

The development of biotechnology should beviewed not so much as the creation of a new in-

du stry as the r evitalization of an old one. Bothfermentation and enzyme technologies willhave an impact on chemical process develop-ment. The first will affect the transition fromnonrenew able to renew able raw m aterials. The

second will allow fermentation-derived prod-ucts to enter the chemical conversion chains,

and will comp ete directly with trad itional chem-ical transformations. (See figure 25. ) Fermenta-

t ion, by replacing various production steps,

could act as a complementary technology in theoverall manufacture of a chemical.



90 q Impacts of Applied Genetics-Micro-Organisms, Plants,and Animals

Figure 25.—Diagram of Alternative Routes to Organic Chemicals

Petroleum Carbohydrates

% followed by number indicatea iength of carbon chain.

SOURCE: (3. E. Tong, “industrial Chemicals From Fermentation Enzymes,” M/croLr. TechrroL, voi. 1,1979, PP. 173-179.

Charac ter is t ics o f b io logical

produc t ion technologies

The major advantages of using commercialfermentation include the use of renewable re-sources, the need for less extreme conditionsdu ring conversion, the use of one-step p rodu c-tion processes, and a reduction in pollution. Amicro-organism might be constructed, for ex-ample, to transform the cellulose in wood di-rectly into ethanol. * (App . 1-D, a case study of the impact of genetics on ethanol production,elaborates these points. )

RENEWABLE RESOURCESGreen plants use the energy captured from

sunlight to transform carbon dioxide from the

“ If IWILI(WI I“OIS iil)~)]S()\’i~l 01 SU(S I 1 i l l l it(s(:oilll)lislllllel~l I)$v I’DNAIcchniqut?s wits sulmlilted to lht> Ik:otl]l)i i l i i i)l DN A A d v i s o r y(kmlnlilltw il l th e %?pl. 25, 1980 mt?eling.

atmosp here into carbohyd rates, some of which

are used for their own energy needs. The restare accumulated in starches, cellulose, lignins,

and other materials called the biomass, wh ich isthe found ation of all renewable resources.

The technologies of genetic engineering couldhelp ease the chemical industry’s dependenceon petroleum-based products by making the useof renewable resources attractive. All micro-organisms can metabolize carbohydrates andconvert them to various end products. Exten-sive research and development (R&D) has

already been conducted on the possibility of using genetically engineered strains to convertcellulose, the major carbohydrate in plants, tocommercial p rodu cts. The basic building block of cellulose—glucose—can be read ily used as araw material for fermentation.



Ch. 5—The Chemical industry q 91

other plant carbohydrates include corn-starch, molasses, and lignin. The last, a polymerfound in w ood, could be used as a precursor forthe biosynthesis of aromatic (benzene-like)chemicals, making their prod uction simp ler andmore economical. Nevertheless, the increase in

the price of petroleum is not a sufficient reasonfor switching raw materials, since the cost of carbohydrates and other biological materialshas been increasing at a relative rate.

PHYSICALLY MILDER CONDITIONS

In general, there are two main ways to speedchemical reactions: by increasing the reactiontemp erature and by add ing a catalyst. A catalyst(usually a m etal or metal complex) causes onespecific reaction to occur at a faster rate thanothers in a chemical mixture by p roviding a sur-

face on which that reaction can be promoted.

Even using the most effective catalyst, the con-ditions needed to accelerate industrial organicreactions often require extremely high tem-peratures and pressures—several hund red d e-grees Celsius and several hund red p ound s persquare inch.

Biological catalysts, or enzymes, on the otherhand can sp eed-up reactions w ithout the needfor such extreme conditions. Reactions occur indilute, aqueous solutions at the mod erate cond i-tions of temperature, pressure, and pH (a meas-ure of the acidity or alkalinity of a solution) that

are compatible with life.ONE-STEP PRODUCTION METHODS

In the chemical synthesis of compounds, each

reaction must take place separately. Becausemost chemical reactions d o not yield pu re prod-ucts, the product of each individual reactionmust be purified before it can be used in thenext step. This approach is time-consuming andexpensive. If, for example, a synthetic schemethat starts with ethylene (a petroleum-basedproduct) requires 10 steps, with each step yield-ing 90 percent product (very optimistic yields inchemical syntheses), only about one-third of theethylene is converted into the final end prod uct.Purification may be costly; often, the chemicalsinvolved (such as organic solvents for extrac-tions) and the byproducts of the reaction aretoxic and require special disposal.

In biological systems, micro-organisms oftencomplete entire synthetic schemes. The conver-sion takes place essentially in a single step,although several might occur within the orga-nisms, whose enzymes can transform the pre-cursor through the intermediates to the desired

end product. Purification is not necessary.

REDUCED POLLUTION

Metal catalysts are often nonspecific in theiraction; while they may promote certain reac-tions, their actions are not or dinar ily limited tomaking only the desired products. Consequent-ly, they have several undesirable features: the

formation of side-products or byproducts; theincomplete conversion of the starting materi-al(s); and the m echanical and accidental loss of the product.

The last problem occurs with all types of syn-thesis. The first two represent inefficiencies inthe u se of the raw materials. These necessitatethe separation and recycling of the side-prod-ucts formed, w hich can be d ifficult and costlybecause they are often chemically and physi-cally similar to th e d esired end prod ucts. (Mostseparation techniques are based on differencesin physical properties—e.g., density, volatility,and size.)

When byproducts and side-products have novalue, or w hen u nconverted raw material can-not be recycled economically, problems of

waste disposal and pollution arise. Their solu-tion requires ingenuity, vigilance, energy, anddollars. Many present chemical processes createuseless wastes that require elaborate degrada-tion procedures to make them environmentallyacceptable. In 1980, the chemical industry is ex-pected to spend $883 million on capital outlaysfor pollution control, and well over $200 million

on R&D for new control techniques and re-placement products. These figures do not in-clude the millions of dollars that have beenspent in recent years to clean up toxic chemicaldumps and to compensate those harmed by

poorly disposed wastes, nor do they include thecost of energy and labor required to operate

pollution-control systems.

A genetically engineered organism, on theother hand, is designed to be precursor- and




product-specific, w i th each enzym e hav ingessentially 100-percent conversion efficiency.An enzymatic process that carries out the same

transformation as a chemical synthesis pro-du ces no side-products (because of an enzyme’shigh sp ecificity to its substrate) or byp rodu cts

(because of an enzyme’s strong catalytic power).Consequently, biological processes eliminatemany conventional waste and disposal problemsat the front end of the system-in the fer-menter. This high conversion efficiency reducesthe costs of recycling. In addition, the efficiencyof the biological conversion process generallysimplifies product recovery, reducing capitaland operating costs. Furthermore, by theirnature, biologically based chemical processes,tend to create some waste products that are bio-degradable and valuable as sources of nutrients.

Specific comparisons of the environmental

hazards produced by conventional and biologi-cal systems are difficult. Data detailing thepollution parameters for various current chem-

ical processes exist, but much less informationis available for fermentation processes, and few

I n d u s t r i a l c h e m i c a l s t h a t

by b io log ica l t e chno log ie s

Despite the benefits of producing industrialchemicals biologically, thus far major fermenta-

tion processes have been developed primarilyfor a few comp lex compoun ds such as enzym es.(See table 11.) Biological methods have also beendeveloped for a few of the simpler commod itychemicals: ethanol, butanol, acetone, aceticacid, isopropanol, glycerol, lactic acid, and citricacid.

Two questions are critical to assessing thefeasibility or d esirability of p rodu cing variouschemicals biologically:

I. Which compounds can be produced bio-

logically (at least theoretically)?2. Which compounds may be primarily de-pendent on genetic technology, given thecosts and availability of raw materials?

compounds are produced by both methods.However, in most beverage distilling operations,pollution has been reduced to almost zero withthe complete recovery of still slops as animalfeeds of high nutritional value. Such controlprocedures are generally applicable to mostfermentation processes. (App. I-C describes thepollutants that may be produced by currentchemical processes and those expected frombiologically based processes.)

The Environmental Protection Agency hasestimated that the U.S. Government and indus-try combined will spend over $360 billion tocontrol air and water pollution in the decadefrom 1977 through 1986. The share of thechemical and allied industries is about $26 bil-lion. Genetic engineering technology m ay h elpalleviate this burden by offering cleaner proc-esses of synthesis and better biological waste

treatment systems. The monetary savings couldbe tremendous. As pure speculation, if just 5percent of the current chemical industry wereaffected, spending on pollution could be re-du ced by abou t $100 million p er year.

m a y b e p r o d u c e d

In principle, virtually all organic compoundscan be produced by biological systems. If the

necessary enzyme or enzymes are not known toexist, a search of the biological world will prob-ably uncover the appropriate ones. Alterna-tively, at least in theory, an enzyme can beengineered to carry out the required reaction.Within this framework, the potential appears tobe limited only by the imagination of the bio-technologist—even though certain chemicalsthat are highly toxic to biological systems areprobably not amenable to prod uction.

Three variables in particular affect theansw er to the second q uestion: the availability

of an organism or enzymes for the desiredtransformation; the cost of the raw material;and the cost of the production process. Whenspecific organisms and production technologies



Ch. 5—The Chemical Industry q 93

Table 1 1.—Some Commercial Enzymes and Their Uses

Enzyme Source Industry and application

Amylase . . . . . . . . . . . . . . . . . . .

Bromelin. . . . . . . . . . . . . . . . . . .

Cellulase and hemicellulase . .Dextransucrase. . . . . . . . . . . . .

Ficin . . . . . . . . . . . . . . . . . . . . . .Glucose oxidase (plus catalase

or peroxidase) . . . . . . . . . . . .

Invertase. . . . . . . . . . . . . . . . . . .

Lactase. . . . . . . . . . . . . . . . . . . .

Lipase. . . . . . . . . . . . . . . . . . . . .Papain . . . . . . . . . . . . . . . . . . . . .

Pectinase . . . . . . . . . . . . . . . . . .Penicillinase . . . . . . . . . . . . . . .

Pepsin . . . . . . . . . . . . . . . . . . . . .Protease. . . . . . . . . . . . . . . . . . .

Streptodornase . . . . . . . . . . . . .

Animal(pancreas)

Plant(barley malt)

Fungi (Aspergillus niger,A. oryzae)

Bacteria (Bacillus subtilis)

Plant (pineapple)

Fungi (Aspergillus niger)Bacteria(Leuconostoc mesenteroides)

Plant (fig latex)

Fungi (Aspergillus niger)

Yeast (Saccharomyces cerevisiae)

Yeast (Saccharomyces fragilis)

Fungi (Aspergillus niger)Plant(papaya)

Fungi (Aspergillus niger)Bacteria(Bacillus cereus)

Animal (hog stomach)Animal (pancreas)

Animal (pepsin)Animal (rennin, rennet)Animal (trypsin)Fungi (Aspergillus oryzae)

Bacteria (Bacillus subtilis)

Bacteria (Streptococcus pyrogenes)

Pharmaceutical digestive aidsTextile: desizing agentBaking: flour supplementBrewing, distilling, and industrial alcohol mashingFood: precooked baby foodsPharmaceutical digestive aidsTextile: desizing agentBaking: flour supplementBrewing, distilling, and industrial alcohol mashingFood: precooked baby foods, syrup manufacturePharmaceutical digestive aidsPaper starch coatingsStarch: cold-swelling laundry starchFood: meat tenderizerPharmaceutical digestive aidsFood: preparation of liquid coffee concentratesPharmaceutical preparation of blood-plasma

extenders, and dextran for other usesPharmaceutical: debriding agent

Pharmaceutical test paper for diabetesFood: glucose removal from egg solids

Candy: prevents granulation of sugars in soft-centercandies

Food: artificial honeyDairy: prevents crystallization of lactose in ice cream

and concentrated milkDairy: flavor production in cheeseBrewing: stabilizes chill-proof beerFood: meat tenderizerWine and fruit juice: clarificationMedicine: treatment of allergic reaction to penicillin,

diagnostic agentFood: animal feed supplementDairy: prevents oxidized flavorFood: protein hydrolysatesLeather: batingPharmaceutical: digestive aidsTextile: desizing agent

Brewing: beer stabilizerDairy: cheesePharmaceutical: wound debridementBaking: breadFood: meat tenderizerBaking: modification of cracker doughBrewing: clarifierPharmaceutical: wound debridement

SOURCE: David Perlman, “The Fermentation Industries,” American Society for Microbiology News 39:10, 1973, p. 653.

have been developed, the cost of raw materials

becomes the limiting step in production. If a

strain of yeast, for example, produces 5 percent

ethanol using sugar as a raw material, the proc-

ess might become economically competitive if th e cost of sugar drops or the price of petro-leum rises. Even if prices remain stable, the

micro-organisms might be genetically improved

to increase their yield; genetic manipulation

m igh t so lve the p rob lem of an ine f f i c i en t

organism. Finally, the p roduction p rocess itself is a factor. After fermentation, the d esired prod -uct must be separated from the other com-pounds in the reaction mixture. As an aid to re-

covery, the production conditions might bealtered and improved to generate more of a de-

sired compound.

More than one raw material can be used in a

fermentation process. If, in the case of ethanol,




the price of sucrose (from sugarcane or sugarbeets) is not expected to change, the productiontechnology is being ru n at optimu m efficiency,and the micro-organism is producing as muchethanol as it can, the hu rd le to econom ic com-petitiveness might be overcome if a less expen-sive raw material—cellulose, perhaps—wereused. But cellulose cannot be u sed in its naturalstate: physical, chemical, or biological methodsmust be devised to break it down to its glucose(also a sugar) components.

The constraints vary from comp ound to com-pound. But even though the role of geneticsmust be examined on a product-by-product ba-sis, certain generalizations can be m ade. Over-all, genetic engineering w ill probably have animpact on three p rocesses:

q

q

q

Aerobic fermentation, which produces en-

zymes, vitamins, pesticides, growth regula-tors, amino acids, nu cleic acids, and otherspeciality chemicals, is already well-estab-lished. Its use should continue to grow. Al-ready, complex biochemical like antibiot-ics, growth factors, and enzymes are madeby fermentation. Amino acids an d nucleo-tides—somewhat less complicated mole-cules—are sometimes produced by fer-mentation. Their production is expected toincrease.Anaerobic fermentation, which producesorganic acids, methane, and solvents, is the

industry’s area of greatest current growth.Already, 40 percent of the ethanol man-ufactured in the United States is producedin this way. The main constraint on theprod uction of other organic acids and sol-vents is the need for cheaper methods forconverting cellulose to fermentable sugars.Chemical modification of the fermentationproducts of both aerobic and anaerobicfermentation, which to date has rarelybeen used on a commercial scale, is of great interest. (See table 12.) Chemical pro-du ction technologies that employ high tem-

peratures and pressures might be replacedby biological technologies operating at at-mospheric pressure and ambient tempera-ture. A patent app lication has alread y beenfiled for the biological production of one of

Table 12.—Expansion of Fermentation Intothe Chemical Industry

Examples

Aerobic fermentationEnzymes . . . . . . . . . . . . . . . . Amylases, proteasesVitamins . . . . . . . . . . . . . . . . Riboflavin B,zPesticides. . . . . . . . . . . . . . . Bacillus thuringiensis

Growth regulators . . . . . . . . GibberellinAmino acids . . . . . . . . . . . . . Glutamic, IysineNucleic acidsAcids. . . . . . . . . . . . . . . . . . . Malic acid, citric acid

Anaerobic fermentationSolvents . . . . . . . . . . . . . . . . Ethanol, acetone, n-butanolAcids. . . . . . . . . . . . . . . . . . . Acetic, propionic, acrylic


these products, ethylene glycol, by theCetus Corp. in Berkeley, Calif. The processis claimed to be m ore energy efficient an dless polluting. If it proves successful whenrun at an industrial scale, the technology

could become significant to a [J. S. markettotaling $21/2 billion per year.

The chemical ind ustry p roduces a variety of likely targets for biotechnology. Tables I-B-27through 1-B-32 in appendix I-B present projec-tions of the potential econom ic impacts of ap-plied genetics on selected compounds thatrepresent large markets, and the time framesfor potential implementation. Table I-B-7 listsone large group of organic chemicals that wereidentified by the Genex Corp. and Massachu-setts Institute of Technology (MIT) as amenable

to biotechnological prod uction method s. Theyare in agreement on about 20 percent of the

products cited, which underscores the uncer-

tain nature of attempting to predict so far intothe future.

Fert i l i zers , polymers , and pes t ic ides

Gaseous ammonia is used to produce nitrogenfertilizers. About 15 billion tonnes of ammoniawere produced chemically for this purpose, in1978; the process requires large amounts of natu ral gas. Nitrogen can also be converted, or

“fixed,” to ammonia by enzymes in micro-orga-nisms; about 175 billion tonnes are fixed peryear. For example, one square yard of landplanted w ith certain legumes (such as soybeans)can fix up to 2 ounces of nitrogen, using bac-



Ch. 5—The Chemical Industry q 95

teria associated with their roots. Currently, mi-crobial prod uction of am monia from nitrogen isnot economically competitive. Aside from thedifficulties associated with the enzyme’s sen-sitivity to oxygen and the near total lack of understanding of its mechanism, it takes the

equivalent of the energy in 4 kilograms (kg) of sugar to make 1 kg of ammonia. Since ammoniacosts $0. 13/ kg and sugar costs $0.22/ kg, it is un -likely that the chem ical process will be rep lacedin the near future. On the other hand, the genesfor nitrogen fixation have now been transferredinto yeast, opening up the possibility that agri-culturally useful nitrogen can be made by fer-mentation.

A large segment of the chemical industry en-gaged in the manufacture of polymers is shownin table 13. A total of 4.3 million tonnes of fibers, 12 million tonnes of p lastics, and I. I mil-lion tonnes of synthetic rubber were p rodu cedin the United States in 1978. All were derivedfrom petroleum, with the exception of the lessthan 1 percent derived from cellulose fibers.The most likely ones are polyam ides (chemicallyrelated to proteins), acrylics, isoprene-type rub-ber, and polystyrene, Because most monomers,the building blocks of polymers, are chemicallysimple and are p resently available in high yieldfrom petroleum, their microbial production inthe n ext decade is u nlikely.

While biotechnoloqy is not r eady to rep lace

the present technology, its eventual impact onpolymer production will probably be large.Biopolymers represent a new w ay of thinking.Most of the imp ortant constituents of cells arepolymers: proteins (polypeptides from aminoacid monomers), polysaccharides (from sugarmonomers), and polynucleotides (from nucleo-tide monomers). Since cells normally assemblepolymers with extreme specificity, the ideal in-dustrial process would imitate the biologicalproduction of polymers in all possible respects—using a single biological machine to convert araw material, e.g., a sugar, into the mon omer to

polymerize it, then to form the final product. Amore likely application is the development of new monomers for specialized applications.Polymer chemistry has largely consisted of thestud y of how their properties can be mod ified.

Table 13.—The Potential of Some Major PolymericMaterials for Production Using Biotechnology

Domestic production 1978Product (thousand tonnes)

PlasticsThermosetting resins

Epoxy. . . . . . . . . . . . . . . . . . . . . . . . .Polyester. . . . . . . . . . . . . . . . . . . . . .Urea. . . . . . . . . . . . . . . . . . . . . . . . . .Melamine . . . . . . . . . . . . . . . . . . . . .Phenolic . . . . . . . . . . . . . . . . . . . . . .

Thermoplastic resinsPolyethylene

Low density . . . . . . . . . . . . . . . . .High density. . . . . . . . . . . . . . . . .

Polypropylene . . . . . . . . . . . . . . . . .Polystyrene. . . . . . . . . . . . . . . . . . . .Polyamide, nylon type. . . . . . . . . . .Polyvinyl alcohol . . . . . . . . . . . . . . .Polyvinyl chloride. . . . . . . . . . . . . . .Other vinyl resins. . . . . . . . . . . . . . .

FibersCellulosic fibers

Acetate . . . . . . . . . . . . . . . . . . . . . . .Rayon . . . . . . . . . . . . . . . . . . . . . . . .

Noncellulosic fibersAcrylic. . . . . . . . . . . . . . . . . . . . . . . .Nylon. . . . . . . . . . . . . . . . . . . . . . . . .Olefins. . . . . . . . . . . . . . . . . . . . . . . .Polyester. . . . . . . . . . . . . . . . . . . . . .Textile glass . . . . . . . . . . . . . . . . . . .Other . . . . . . . . . . . . . . . . . . . . . . . . .

RubbersStyrene-butadiene. . . . . . . . . . . . . . . .Polybutadiene . . . . . . . . . . . . . . . . . . .Butyl . . . . . . . . . . . . . . . . . . . . . . . . . . .Nitrile. . . . . . . . . . . . . . . . . . . . . . . . . . .Polychlorophene . . . . . . . . . . . . . . . . .Ethylene-propylene . . . . . . . . . . . . . . .Polyisoprene. . . . . . . . . . . . . . . . . . . . .

135

544504

90727

3,2001,8901,3802,680

12457

2,57588

139269

327 1,148

3111,710

4187

6281706933727862


Conceivably, biotechnology could enable themodification of their function and form.

Pesticides include fungicides, herbicides, in-secticides, rodenticides, and related productssuch as plant growth regu lators, seed d isinfec-tants, soil conditioners, and soil fumigants. Thelargest market (roughly $500 million a nnu ally)involves the chemical and microbial control of insects. Although microbial insecticides havebeen around for years, they comprise only 5

percent of the market. However, recent suc-cesses in developing viruses and bacteria thatproduce diseases in insects, and the negativepublicity given to chemical insecticides, haveencouraged the use of microbial insecticides.




The other two species can be produced by con-Ventional fermentation techniques. They havebeen useful because they form spores that canbe mass-produced easily and are stable enoughto be handled commercially. The actual sub-stances that cause toxicity to the insect are tox-

ins synthesized by the microbes.Genetic engineering should make it p ossible

to construct m ore p otent bacterial insecticidesby increasing the dosage of the genes that codefor the synthesis of the toxins involved. Mix-tures of gen es capable of directing the synth esisof various toxins might also be prod uced.

C o n s t r a i n t s o n b i o lo g ic a l p r o d u c t i on t e c h n i q u e s ____

Th e chief impediments to using biological

production technology are associated with theneed for biomass.

2They include:

q

q

q

q

q

q

q

competition with food n eeds for starch andsugar;cyclic availability;biodegradability and associated storageproblems;high moisture content for cellulosics, and

high collection and storage costs;mechanical processing for cellulosics;the heterogeneous nature of cellulosics (mix-tures of cellulose, hemicellulose, and lig-nin); andThe need for disposal of the nonferment-able port ions of the biomass.

For food-related b iomass sources, such as su-gar, corn, and sorgh um , few technological bar-riers exist for conversion to fermentable sugars;but subsidies are needed to make the fermenta-tion of sugars as profitable as their use as food.For cellulosic biomass sources such as agricul-tural wastes, municipal wastes, and wood, tech-nological barriers exist in collection, storage,pretreatment, fermentation, and waste disposal.In addition, biomass must always be trans-

formed into sugars by either chemical or en-zymatic processes before fermentation canbegin.

‘ljnf~r,qv F’r(ml ~ io / (J , g i f ’ i f / PrOct’s.st*,s, 01). (’it.

A second major impediment is associated

with th e pu rification stage of prod uction. Mostchemical products of fermentation are presentin extremely dilute solutions, and concentratingthese solutions to recover the desired product ishighly energy-intensive. Problems of technologyand cost will continu e to make this stage an im-portant one to improve.

The developments in genetics show greatprom ise for creating m ore versatile micro-orga-nisms, but they do not by themselves produce acheaper fuel or plastic. Associated technologiesstill require more efficient fermentation facil-ities and product separation processes; mi-

crobes may produce molecules, but they willnot isolate, purify, concentrate, mix, or packagethem for hu man u se.

The interaction betw een genetic engineeringand other technologies is illustrated by theproblems of producing ethanol by fermenta-tion. The case stud y presented in ap pend ix I-Didentifies those steps in the biomass-to-ethanolscheme that need technological improvementsbefore the process can become economical.

Genetic engineering is expected to reducecosts in many production steps. For certainones—such as the pretreatm ent of the biomassto make it fermentable—genetics will probablynot p lay a role: physical and chemical technol-ogies will be responsible for the greatest ad-vances. For others, such as distillation, genetic




Table 14.—Some Private Companies With Biotechnology Programs

Approximate Research capacity

Company Founded employees 1979 Ph. D.s 1979 Recombinant DNA Hybridomas

Atlantic Antibodies. . . . . . . . . . . . . . . . . . 1973 50 2 xBethesda Research Laboratories . . . . . . 1976 130 x xBiogen . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1978 30 (50b) (18

b)

(3)(5)

x x

Bens Bio Logicals . . . . . . . . . . . . . . . . . . . 1979 15 10 x xCentocor. . . . . . . . . . . . . . . . . . . . . . . . . . . 1979 20(1)-10(4) xCetus . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1972 250 50 x xClonal Research . . . . . . . . . . . . . . . . . . . . 1979 6 1 xCollaborative Research

C. . . . . . . . . . . . . . 1961 85 15 x x

(Collaborative Genetics) . . . . . . . . . . . . (1979) (4) (3) xGenentech . . . . . . . . . . . . . . . . . . . . . . . . . 1976 90 30 xGenex . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1977 30 12 xHybritech . . . . . . . . . . . . . . . . . . . . . . . . . . 1978 33(1)

6 xMolecular Genetics. . . . . . . . . . . . . . . . . . 1979 6(4) x xMonoclinal Antibodies. . . . . . . . . . . . . . . 1979 6 xNewEngland Biolabs . . . . . . . . . . . . . . . . 1974 22(22)-5 (4) x

q=.E~r9t~t~co. estimates.bEXwtedby~cem~rl~.ccollaboratlve Research isamajorowner of Collaborative Genetics. Thedivision batweenthem isnotyetdistinct.

SOURCES: (l)Sc/ence206, p.692.693, 1960(52 peopletoexpandto100 by1961).(2) Science 206, p. 692-693, 19S0 (20 senior peraona).

(3) Science 206, p. 692-693, 1960(16 scientists, 30 employees).

(4 ) Dun & Bradstreet, Inc.

(5) Chemlcalarrd Engineering News, Mar. 19,1960.Office of Technology Assessment.

neering firm also intends to man ufacture someprod ucts itself. It is likely that the p rodu cts re-served for inhouse manufacture will be low-volume, high-priced compoun ds like interferon.

Genetic engineering by itself is a relativelysmall-scale laboratory operation. Consequently,genetic engineering firms will continue to offerservices to companies that do not intend todevelop th is capacity in their own inh ouse lab-

oratories. Specifically, a genetic engineeringcompany may contract with a firm to develop abiological production method for its products.At the sam e time, larger compan ies might estab-lish inhou se staffs to develop biological meth odsfor both old and new prod ucts. (Several largercompanies already have more inhouse geneticengineering personnel than some of the inde-pend ent genetic engineering comp anies.)

In ad dition, suppliers of genetic raw materialsmay decide to expand into the production of genetically engineered orga nisms. Sup pliers of

restriction endonuclease enzymes for example,which are used in constructing rDNA, havealready entered the field. Diagnostic firms coulddevelop new bioassays for which they them-selves would guarantee a market. Finally, com-panies with byprodu cts or waste produ cts are

beginning to examine the possibility of convert-ing them into useful products. This approach(which is somewhat more developed in Europe)assumes that with the proper technology thewaste m aterials can become a resource.

Some ind ustries, includ ing man ufacturers of agitators (drives), centrifuges, evaporators, fer-menters, dryers, storage tanks and processvessels, and control and instrumentation sys-tems, might profit by producing equipmentassociated w ith fermentation.

Impac ts on univers i ty research

From the beginning, genetic engineeringfirms established stron g ties with un iversities.These were responsible for providing most of the scientific knowledge th at formed the basisfor applied genetics as well as the initial scien-tific workforce:

Cetus Corp. established a pattern by re-cruiting a prestigious Board of ScientificAdvisors who remain in academic posi-tions.

Genentech, Inc., cofoun ded by a professorat the University of California at San Fran-



100 . Impacts of Applied Genetics—Micro-0rganisms, Plants, and Animals

Data obtained th rough an (OTA survey of 284firms indicate that the pharmaceutical industryemp loys the major share of personnel workingin applied genetics programs. (See table 15. ) Theaverage number of Ph. D.s in each industry isgiven in table 16. A rough estimate of profes-sional scientific manpow er at this level includ es:6 in food, 45 in chemical, 120 in ph arm aceutical,and 18 in specialty chemicals-a total of 189. If the number of research support personnel isapproximately twice the number of Ph. D.s, thetotal rises to about 570. If $165,000 per year isrequired to sup port one Ph. D. in industry, thetotal value of such manpower is approximately

$31 million.

Estimates of the number of companies en-gaged in applied genetics work in 1980 can becomp ared w ith the total num ber of firms withfermentation activities. A tabulation of firms on

a worldwide basis in 1977 revealed 145 com-panies, of which 27 were American. (See table17. ) These companies produced antibiotics, en-zymes, solvents, vitamins and growth factors,

Table 15.—Distribution of Applied GeneticsActivity in Industry

Distribution of appliedgenetics activity by Percent

Classification company classa

of total

Food . . . . . . . . . . . . . . (6146) 13Chemical . . . . . . . . . . (9/52) 17

Pharmaceutical . . . . . (12/25) 48Specialty chemicalb . (6/68) 9

algnoreg small firing specializing in genetic research.bFood ingredients, reagents, enzymes.


Table 16.—Manpower (Iow.(average)-high) Distributionof a Firm With Applied Genetics Activity

Ph. D. M.S. Bachelors

Food. . . . . . . . . . . . . . . . . ql)-2Chemical . . . . . . . . . . . . . 3-(5)-7Pharmaceutical. . . . . . . 2-(10)-24Specialty. . . . . . . . . . . . . l-(3)-8

BiotechnologyGenetic engineering. 3-(15)-32Hybridoma. . . . . . . . . . l-(3)-6Other . . . . . . . . . . . . . . O-(2)-4

Average . . . . . . . . . .1-(6)-12

0-(1 )-20-(1 )-2l-(4)-9l-(3)-4

2-(1 1)-200-(2)-02-(4)-6l-(4)-6

O-(2)-82-(5)-71-(8)-202-(2)-4

5-(15)-250-(20)-0

8-(10)-133-(8)-12


Table 17.—lndex to Fermentation Companies

1. Abbott Laboratories, North Chicago, 111.

2. American Cyanamid, Wayne, N.J.3. Anheuser-Busch, Inc., St. Louis, Mo.4. Bristol-Myers Co., Syracuse, N.Y.5. Clinton Corn Processing Co., Clinton, Iowa6. CPC International, Inc., Argo, Ill.7. Dairyland Laboratories, Inc., Waukesha, Wis.

8. Dawe’s Laboratories, Inc., Chicago Heights, Ill.9. Grain Processing Corp., Muscatine, Iowa

10. Hoffman-LaRoche, Inc., Nutley, N.J.11. IMC Chemical Group, Inc., Terre Haute, Ind.12. Eli Lilly & Co., Indianapolis, Ind.13. Merck & Co., Inc., Rahway, N.J.14. Miles Laboratories, Inc., Elkhart, Ind.15. Parke, Davis & Co., Detroit, Mich.16. S. B. Penick & Co., Lyndhurst, N.J.17. Pfizer, Inc., New York, N.Y.18. Premier Malt Products, Inc., Milwaukee, Wis.19. Rachelle Laboratories, Inc., Long Beach, Cal if.20. Rohm & Haas, Philadelphia, Pa.21. Schering Corp., Bloomfield, N.J.22. G. D. Searle & Co., Skokie, Ill.23. E. R. Squibb& Sons, Inc., Princeton, N.J.24. Standard Brands, Inc., New York, N.Y.

25. Stauffer Chemical Co., Westport, Corm.26. Universal Foods Corp., Milwaukee, Wis.27. The Upjohn Co., Kalamazoo, Mich.28. Wallerstein Laboratories, Inc., Morton Grove, Ill.29. Wyeth Laboratories, Philadelphia, Pa.


nucleosides, amino acids, and miscellaneousproducts. (See table 18.) The only chemical firm

listed was the Stauffer Chemical Co. Ten firms

are listed as having the ability to produce foodand feed yeast. (See table 19. ) Correcting forfirms listed twice, at least 38 U.S. firms were

engaged in significant fermentation activity forcommercial products, excluding alcoholic bev-erages, in 1977. Not all have research expertisein fermentation or biotechnology, mu ch less aregular gen etics prog ram: 10 to 20 were in thechemical industry; 25 to 40 in fermentation (en-zyme, pharmaceutical, food, and specializedchemicals); and 10 to 15 in biotechnology (genet-ic engineering)—or abou t 45 to 75 firms in all.

If average manpow er num bers are used, thetotal num ber of professionals involved in com-mercial applied genetics research is:

Ph. D.s: 300-450

Others: 600-900900-1,350

The number of workers that will be involvedin the p rodu ction p hase of biotechnology repre-



Ch.5—The Chemical Industry q 101

Table 18.-Fermentation Products and Producers

Product Some producers Product Some producers

Amino acidsL-alanine. . . . . . . . . . . . . . . . . . . . . . . . . . .L-arginine. . . . . . . . . . . . . . . . . . . . . . . . . .L-aspartic acid. . . . . . . . . . . . . . . . . . . . . .

L-citrulline . . . . . . . . . . . . . . . . . . . . . . . . .L-glutarnic acid . . . . . . . . . . . . . . . . . . . . .L-glutamine . . . . . . . . . . . . . . . . . . . . . . . .L-glutathione . . . . . . . . . . . . . . . . . . . . . . .L-histidine . . . . . . . . . . . . . . . . . . . . . . . . .L-homoserine. . . . . . . . . . . . . . . . . . . . . . .L-isoleucine . . . . . . . . . . . . . . . . . . . . . . . .L-leucine. . . . . . . . . . . . . . . . . . . . . . . . . . .L-lysine . . . . . . . . . . . . . . . . . . . . . . . . . . . .L-methionine . . . . . . . . . . . . . . . . . . . . . . .L-ornithine . . . . . . . . . . . . . . . . . . . . . . . . .L-phenylalanine. . . . . . . . . . . . . . . . . . . . .L-proline. .... . . . . . . . . . . . . . . . . . . . . .L-serine. . . . . . . . . . . . . . . . . . . . . . . . . . . .L-threonine. .. . . . . . . . . . . . . . . . . . . . . .L-tryptophan. . . . . . . . . . . . . . . . . . . . . . .L-tyrosine . . . . . . . . . . . . . . . . . . . . . . . . . .

L-valine. ..... . . . . . . . . . . . . . . . . . . . . .Miscellaneous products and processesAcetoin . . . . . . . . . . . . . . . . . . . . . . . . . . . .Acyloin . . . . . . . . . . . . . . . . . . . . . . . . . . . .Anka-pigment(red) . . . . . . . . . . . . . . . . . .Blue cheese flavor. . . . . . . . . . . . . . . . . . .Desferrioxamine . . . . . . . . . . . . . . . . . . . .Dihydroxyacetone . . . . . . . . . . . . . . . . . . .Dextran . . . . . . . . . . . . . . . . . . . . . . . . . . . .Diacetyl (from acetoin) . . . . . . . . . . . . . . .Ergocornine . . . . . . . . . . . . . . . . . . . . . . . .Ergocristine . . . . . . . . . . . . . . . . . . . . . . . .Ergocryptine . . . . . . . . . . . . . . . . . . . . . . .Ergometrine . . . . . . . . . . . . . . . . . . . . . . . .Ergotamine. . . . . . . . . . . . . . . . . . . . . . . . .Bacillus thuringiensis insecticide. . . . . .Lysergic acid . . . . . . . . . . . . . . . . . . . . . . .

Paspalic acid . . . . . . . . . . . . . . . . . . . . . . .Picibanil . . . . . . . . . . . . . . . . . . . . . . . . . . .Ribose. . . . . . . . . . . . . . . . . . . . . . . . . . . . .Scteroglucan . . . . . . . . . . . . . . . . . . . . . . .Sorbose(from sorbitol). . . . . . . . . . . . . . .Starter cultures . . . . . . . . . . . . . . . . . . . . .Sterol oxidations.. . . . . . . . . . . . . . . . . . .Steroid oxidations. . . . . . . . . . . . . . . . . . .Xanthan . . . . . . . . . . . . . . . . . . . . . . . . . . .

AntibioticsAdriamycin. . . . . . . . . . . . . . . . . . . . . . . . .Amphomycin . . . . . . . . . . . . . . . . . . . . . . .Amphotericin B.. . . . . . . . . . . . . . . . . . . .Avoparcin . . . . . . . . . . . . . . . . . . . . . . . . . .Azalomycin F . . . . . . . . . . . . . . . . . . . . . . .Bacitracin. . . . . . . . . . . . . . . . . . . . . . . . . .Bambermycins. . . . . . . . . . . . . . . . . . . . . .

Bicyclomycin. . . . . . . . . . . . . . . . . . . . . . .Blasticidin S. . . . . . . . . . . . . . . . . . . . . . . .Bleomycin . . . . . . . . . . . . . . . . . . . . . . . . .CactinomycinCandicidin B. . . . . . . . . . . . . . . . . . . . . . . .Candidin . . . . . . . . . . . . . . . . . . . . . . . . . . .

25

13

7

17,21,28

1

10,177,13,1422,2721,23,27,2913,17

232

11,16,17

16

Capreomycin . . . . . . . . . . . . . . . . . . . . . . .Cephalosporins. . . . . . . . . . . . . . . . . . . . .Chromomycin A3 . . . . . . . . . . . . . . . . . . . .Colistin . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Cycloheximide. . . . . . . . . . . . . . . . . . . . . .Cycloserine . . . . . . . . . . . . . . . . . . . . . . . .Dactinomycin. . . . . . . . . . . . . . . . . . . . . . .Daunorubicin. . . . . . . . . . . . . . . . . . . . . . .Destomycin . . . . . . . . . . . . . . . . . . . . . . . .Enduracidin . . . . . . . . . . . . . . . . . . . . . . . .Erythromycin . . . . . . . . . . . . . . . . . . . . . . .Fortimicins. . . . . . . . . . . . . . . . . . . . . . . . .Fumagiliin . . . . . . . . . . . . . . . . . . . . . . . . .Fungimycin . . . . . . . . . . . . . . . . . . . . . . . .Fusidic acid . . . . . . . . . . . . . . . . . . . . . . . .Gentamicins. . . . . . . . . . . . . . . . . . . . . . . .Gramicidin A . . . . . . . . . . . . . . . . . . . . . . .Gramicidin J(S) . . . . . . . . . . . . . . . . . . . . .Griseofulvin . . . . . . . . . . . . . . . . . . . . . . . .Hygromycin B . . . . . . . . . . . . . . . . . . . . . .Josamycin . . . . . . . . . . . . . . . . . . . . . . . . .Kanamycins. . . . . . . . . . . . . . . . . . . . . . . .

Kasugamycin. . . . . . . . . . . . . . . . . . . . . . .Kitasatamycin. . . . . . . . . . . . . . . . . . . . . .Lasalocid . . . . . . . . . . . . . . . . . . . . . . . . . .Lincomycin. . . . . . . . . . . . . . . . . . . . . . . . .Lividomycin . . . . . . . . . . . . . . . . . . . . . . . .Macarbomycin. . . . . . . . . . . . . . . . . . . . . .Mepartricin. . . . . . . . . . . . . . . . . . . . . . . . .Midecamycin. . . . . . . . . . . . . . . . . . . . . . .Mikamycins . . . . . . . . . . . . . . . . . . . . . . . .Mithramycin. . . . . . . . . . . . . . . . . . . . . . . .Mitomycin C. . . . . . . . . . . . . . . . . . . . . . . .Mocimycin . . . . . . . . . . . . . . . . . . . . . . . . .Monensin . . . . . . . . . . . . . . . . . . . . . . . . . .Myxin . . . . . . . . . . . . . . . . . . . . . . . . . . . . .Neornycins. ... . . . . . . . . . . . . . . . . . . . .Novobiocin. . . . . . . . . . . . . . . . . . . . . . . . .Nystatin . . . . . . . . . . . . . . . . . . . . . . . . . . .

Oleandomycin. . . . . . . . . . . . . . . . . . . . . .Oligomycin. . . . . . . . . . . . . . . . . . . . . . . . .Paromomycins. . . . . . . . . . . . . . . . . . . . . .Penicillin G. . . . . . . . . . . . . . . . . . . . . . . . .Penicillin V . . . . . . . . . . . . . . . . . . . . . . . . .Penicillins(semisynthetic). . . . . . . . . . . .Pentamycin . . . . . . . . . . . . . . . . . . . . . . . .Pimaricin . . . . . . . . . . . . . . . . . . . . . . . . . .Polymyxins. . . . . . . . . . . . . . . . . . . . . . . . .Polyoxins . . . . . . . . . . . . . . . . . . . . . . . . . .Pristinamycins. . . . . . . . . . . . . . . . . . . . . .Quebemycin. . . . . . . . . . . . . . . . . . . . . . . .Ribostamycin. . . . . . . . . . . . . . . . . . . . . . .Rifamycins. . . . . . . . . . . . . . . . . . . . . . . . .Sagamicin. . . . . . . . . . . . . . . . . . . . . . . . . .Salinomycin, . . . . . . . . . . . . . . . . . . . . . . .Siccanin . . . . . . . . . . . . . . . . . . . . . . . . . . .

Siomycin. . . . . . . . . . . . . . . . . . . . . . . . . . .Sisomicin ... . . . . . . . . . . . . . . . . . . . .Spectinomycin. . . . . . . . . . . . . . . . . . . . . .Streptomycin. . . . . . . . . . . . . . . . . . . . . .

TetracyclinesClortetracycline. . . . . . . . . . . . . . . . . . . . .

4,12

271113

17,27

2128

12

4

1027

174

1016,17,23,272723

17

154,12,13,17,23,291,4,12,17,23,294,13,17,23,29

17

212713,17,29

19

76-565 0 - 81 - 8



102 . Impacts of Applied Genetics—Micro-Organkms, Plants, and Animals

Table 18.-Fermentation Products and Producers

Product Some producers Product Some producers

Demeclocycline. . . . . . . . . . . . . . . . . . . . .Oxytetracycline. . . . . . . . . . . . . . . . . . . . .Tetracycline. . . . . . . . . . . . . . . . . . . . . . . .Tetranactin. . . . . . . . . . . . . . . . . . . . . . . . .

Thiopeptin . . . . . . . . . . . . . . . . . . . . . . . . .Thiostrepton . . . . . . . . . . . . . . . . . . . . . . .Tobramycin . . . . . . . . . . . . . . . . . . . . . . . .Trichomycin. . . . . . . . . . . . . . . . . . . . . . . .Tylosin . . . . . . . . . . . . . . . . . . . . . . . . . . . .Tyrothricin ..... . . . . . . . . . . . . . . . . . . .Tyrocidine . . . . . . . . . . . . . . . . . . . . . . . . .Uromycin . . . . . . . . . . . . . . . . . . . . . . . . . .Vaildamycin . . . . . . . . . . . . . . . . . . . . . . . .Vancomycin . . . . . . . . . . . . . . . . . . . . . . . .Variotin. . . . . . . . . . . . . . . . . . . . . . . . . . . .Viomycin. .. ..... . . . . . . . . . . . . . . . . .Virginiamycin. . . . . . . . . . . . . . . . . . . . . .

EnzymesAmylases . . . . . . . . . . . . . . . . . . . . . . . . . .Amyloglucosidase. . . . . . . . . . . . . . . . . . .Anticyanase. . . . . . . . . . . . . . . . . . . . . . . .

L-asparaginase . . . . . . . . . . . . . . . . . . . . .Catalase . . . . . . . . . . . . . . . . . . . . . . . . . . .Cellulase. . . . . . . . . . . . . . . . . . . . . . . . . . .Dextranase. . . . . . . . . . . . . . . . . . . . . . . . .‘Diagnostic enzymes’ . . . . . . . . . . . . . . . .Esterase-lipase . . . . . . . . . . . . . . . . . . . . .Glucanase . . . . . . . . . . . . . . . . . . . . . . . . .Glucose dehydrogenase. . . . . . . . . . . . . .Glucose isomerase . . . . . . . . . . . . . . . . . .Glucose oxidase . . . . . . . . . . . . . . . . . . . .Glutamic decarboxylase. . . . . . . . . . . . . .Hemi-celiulase. . . . . . . . . . . . . . . . . . . . . .Hespiriginase. . . . . . . . . . . . . . . . . . . . . . .lnvertase. . . . . . . . . . . . . . . . . . . . . . . . . . .

217,192,4,17,19,23,27

2312

1216,28

12

5,19,20,24,285,6,14,28

8,146,20,28

2828

3,5,14,248,141814,20,28

24,26,28

Lactase . . . . . . . . . . . . . . . . . . . . . . . . . . . .Lipase . . . . . . . . . . . . . . . . . . . . . . . . . . . . .Microbial rennet. . . . . . . . . . . . . . . . . . . . .Naringinase . . . . . . . . . . . . . . . . . . . . . . . .

Pectinase . . . . . . . . . . . . . . . . . . . . . . . . . .Pentosanase . . . . . . . . . . . . . . . . . . . . . . .Proteases . . . . . . . . . . . . . . . . . . . . . . . . . .Streptokinase-streptodornase. . . . . . . . .Uricase . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Organic acidsCitric acid . . . . . . . . . . . . . . . . . . . . . . . . . .Comenic acid. . . . . . . . . . . . . . . . . . . . . . .Erythorbic acid. . . . . . . . . . . . . . . . . . . . . .Gluconic acid. . . . . . . . . . . . . . . . . . . . . . .Itaconic acid. . . . . . . . . . . . . . . . . . . . . . . .2-keto-D-giuconic acid . . . . . . . . . . . . . . .~-ketoglutaric acid. . . . . . . . . . . . . . . . . . .Lactic acid . . . . . . . . . . . . . . . . . . . . . . . . .Malic acid . . . . . . . . . . . . . . . . . . . . . . . . . .Urocanic acid. . . . . . . . . . . . . . . . . . . . . . .

SolventsEthanol . . . . . . . . . . . . . . . . . . . . . . . . . . . .2,3-butanediol . . . . . . . . . . . . . . . . . . . . . .

Vitamins and growth factorsGibbereliins . . . . . . . . . . . . . . . . . . . . . . . .Riboflavin . . . . . . . . . . . . . . . . . . . . . . . . . .Vitamin B12. . . . . . . . . . . . . . . . . . . . . . . . .Zearalanol. . . . . . . . . . . . . . . . . . . . . . . . . .

Nucleosides and nucleotides5-ribonucieotides and nucleosides . . . . .Orotic acid . . . . . . . . . . . . . . . . . . . . . . . . .Ara-A-(9-B-D-arabino-furanosyl) . . . . . . ..

6-azauridine . . . . . . . . . . . . . . . . . . . . . . . .

282017,2828

20,2820,2814,17,18,20,282

14,1717

4,17,181717

5

9

1,12,13131311

15

aBlankm=n~ noU.S.pr~ucer in 1977;therefore, l~produeedbyoneormore foreign flrms(fromatleast 120different firms).

SOURCE: OtticeotTechnology Assessment.

sents a major impact of genetic engineering. Toestimate this number these two calculationsmust be made:

q the value or volume of chemicals thatmight be prod uced by fermentation, and

q the num ber of prod uction w orkers neededper u nit volum e of chemicals prod uced.

Any prediction of the potential volume of chemicals is necessarily filled with uncertain-

ties. The app roximate market valu e of organicchemicals produced in the United States is givenin appendix I-B. Total U.S. sales in 1979 werecalculated to be over $42 billion. On the basis of the assumptions made, $522 million worth of bulk organic chemicals could be comm ercially

produced by genetically engineered strains in10 years and $7.1 billion in 20 years. TableI-B-l0 in ap pen dix I-B lists the potential marketsfor pharmaceuticals. Excluding methane pro-duction, the total potential market for productsobtained from genetically engineered orga-nisms is approximately $14.6 billion.

If the production of chemicals having thisvalue is carried out by fermentation, it impossi-ble to calculate how many workers will beneeded. Data obtained from industrial sourcesreveal that 2 to 5 workers, including those insupervision, services, and production, are re-quired for $l million worth of produ ct. Hence,30,000 to 75,000 workers would be required forthe estimated $14.6 billion market.



chapter 6

The Food Processing Industry



Ch a p t e r 6

Page

Introd uction —The Ind ustry . . . . . . . . . . . . . .. .107Single-Cell Protein. . . . . . . . . . . . . . . . . . . . . .. .107

Genetic Engineering and SCP Produ ction. ... 109Commercial Production. . ................109

Genetics in Backing, Brewing, and Winemaking .110Microbial Polysaccharides. . , . . . . . . . . . . . . . . .111Enzymes. . . . . . . . . . . . . . . . . . . . . , . ........111

Genetic Engineering and Enzymes in theFood Pr ocessin g In du stry . . . . . . . . . . . . . . . 112

Sweeteners, Flavors, and Fragrances . . . . . . . . 112Overview . . . . . . . . . . . . . . . . . . . . ..........113

Tables

21. Classification of Yeast-Related U.S. Patents , , 10822. Comparison of Selling Price Ranges for

Selected Microbial, Plant, and AnimalProtein Products. . . . . . . . . . ............108

23, Raw Materials Already Tested on aLaboratory or Small Plant Scale. ... , , .. ...109

Figure

Figure No. Page

26. The Use of Hybridization To Obtain a YeastStrain for the Production of Low-Carbohydrate Beer . . . . . . . . . . . . ........110

Table No. Page

20. Estimated Annual Yeast Production, 1977...108



Chapter 6

T h e F o o d P r o c e s s i n g I n d u s t r y

I n t r o d u c t i o n — t h e i n d u s t r y

The food processing industry comprisesthose manufacturers that transform or process

agricultural products into edible products formarket. It is distinguished from the p rodu ction,or farming and breeding portions of the agricul-tural industry.

Genetics can be used in the food processingindustry in two ways: to design micro-orga-nisms that transform inedible biomass into foodfor human consump tion or for animal feed; andto design organisms that aid in food p rocessing,either by acting directly on the food itseIf or byproviding materials that can be added to food.

Eight million to ten million people work in the

meat, poultry, dairy, and baking industries; incanned, cured, and frozen food plants; and inmoving food from the farm to the dinner table.In 1979, the payroll was over $3.2 billion for t hemeat and poultry industries, $2.6 billion forbaking, an d $1.9 billion for food p rocessing.

Single-ce l l p ro te in

The interest in augmenting the world’s sup-ply of protein has focused attention on micro-

bial sources of protein as food for both animalsand humans. * Since a large portion of eachbacterial or yeast cell consists of proteins (up to72 percent for some protein-rich cells), largenumbers have been grown to supply single-cellprotein (SCP) for consumption, The protein canbe consumed directly as part of the cell itself orcan be extracted and processed into fibers ormeat-like items. By now, ad vanced food proc-essing technologies can combine this proteinwith meat flavoring and other substances toproduce nutritious food that looks, feels, andtastes like meat.

*AS an example of th e potential significance of SCP, the SovietI Inion, which is one of the largest producers, expects to produceenough fodder yeast from in ternally available raw m aterials to beself-sufficient in animal protein foodstuffs by 1990.

Traditionally, micro-organisms have beenused to stabilize, flavor, and modify various

prop erties of food. More recently, efforts havebeen made to control microbial spoilage and toensure that foods are free from micro-orga-nisms that may be hazardous to pu blic health.These are the tw o major ways in w hich micro-biology has been useful.

Historically, most efforts hav e been d evotedto improv ing the ability to control the harm fuleffects of micro-organisms. The industry recog-nized the extreme heat resistance of bacterialspores in the early 20th centu ry and sponsoredor conducted much of the early research on themechanisms of bacterial spore heat resistance.

Efforts to exploit the beneficial characteristicsof micro-organisms, on the other hand, havebeen largely through trial-and-error. Strainsthat improve the quality or character of foodgenerally have been found, rather than de-signed.

The idea of using SCP as animal feed orhu man food is not new ; yeast has been used as

food protein since the beginning of the century.How ever, in th e past 15 years, there has been adram atic increase in research on SCP and in theconstruction of large-scale plants for its pro-du ction, especially for the prod uction of yeast.(See table 20.) Interest in this material is re-flected in the numerous national and interna-tional conferences on SCP, the increasingnumber of proceedings and reviews published,and the number of patents issued in recent

years. (See table 21.)

The issues addressed have covered topics

such as the economic and technological factorsinfluencing SCP processes, nutrition and safety,and SCP app lications to hum an or an imal foods.Thus far, commercial use has been limited by

107




Table 20.-Estimated Annual Yeast Production, 1977(dry tonnes)

Baker’s yeast Dried yeasta

Europe. . . . . . . . . . . . . . . . . . 74,000 b160,000

b

North America . . . . . . . . . . . 73,000 53,000The Orient. . . . . . . . . . . . . . . 15,000 25,000United Kingdom. . . . . . . . . . 15,500 (c)South America. . . . . . . . . . . 7,500 2,000Africa. . . . . . . . . . . . . . . . . . . 2,700 2,500

Totals . . . . . . . . . . . . . . . . 187,700 242,500

aDrigd yeast includes food and fodder yeasts; data for petroleum-grown Yeasts

are not available.bproduction figures for U.S.S.R. not reported.cNone reported.

SOURCE: H. J. Peppier and D. Perlman (eds.), Microbial Technology, vol. 1 (Lon-don: Academic Press, 1979), p. 159.

Table 21.—Classification of Yeast-RelatedU.S. patents (1970 to July 1977)

Category Number issued

Yeast technology (apparatus, processing) . . . . 22

Growth on hydrocarbons. ., . . . . . . . . . . . . . . . . 28Growth on alcohols, acids, wastes. . . . . . . . . . . 22Production of chemicals. . . . . . . . . . . . . . . . . . . 14Use of baking and pasta products . . . . . . . . . . . 24Condiments and flavor enhancers . . . . . . . . . . . 18Reduced RNA. . . . . . . . . . . . . . . . . . . . . , . . . . . . 11Yeast modification of food products . . . . . . . . . 13Isolated protein . . . . . . . . . . . . . . . . . . . . . . . . . . 5Texturized yeast protein . . . . . . . . . . . . . . . . . . . 7Lysates and ruptured cells . . . . . . . . . . . . . . . . . 7Animal feed supplements . . . . . . . . . . . . . . . . . . 12

Total. ., . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 183

SOURCE: H. J. Peppier, “Yeast,” Annual Report on Fermentation Processes, D.Perlman (cd.), vol. 2 (London: Academic Press, 1978), pp. 191-200.

several factors. For each bacterial, yeast, or

algal strain used , technological p roblems (fromthe choice of micro-organisms to the use of cor-responding raw material) and logistical prob-lems of construction and location of plants havearisen. But the primary limitation so far hasbeen the cost of produ ction comp ared w ith thecosts of comp eting sources of protein. (The com-parative price ranges in 1979 for selectedmicrobial, plant, and animal protein productsare shown in table 22.)

The costs of manufacturing SCP for animalfeed in the United States are high, pa rticularlyrelative to its major competing p rotein source,soybeans, which can be p rodu ced w ith little fer-tilizer and minimal p rocessing. The easy avail-ability of this legume severely limits microbialSCP prod uction for animal feed or h um an food.In fact, according to the U.S. Department of

Table 22.—Comparison of Selling Price Rangesfor Selected Microbial, Plant, and Animal

Protein Products

Crude Price rangeprotein 1979 Us.

Product, substrate, and quality content dollars/kg

Single cell proteinsCandida utilis, ethanol, food grade 52 1.32-1.35Kluyveromyces tragilis, cheese

whey, food grade. . . . . . . . . . . . . 54 1.32Saccharomyces cerevisiae:

Brewer’s, debittered, food grade 52 1.00-1.20Feed grade. . . . . . . . . . . . . . . . . . 52 0.39-0.50

Plant proteinsAlfalfa (dehydrated) . . . . . . . . . . . . 0.12-0.13Soybean meal, defatted. . . . . . . . . 0.20-0.22Soy protein concentrate. . . . . . . . . 70-72 0.90-1.14Soy protein isolate . . . . . . . . . . . . . 90“ 92 1.96-2.20

Animal proteinsFishmeal (Peruvian) . . . . . . . . . . . . 65 0.41-0.45Meat and bonemeal . . . . . . . . . . . . 50 0.24-0.25Dry skim milk. . . . . . . . . . . . . . . . . . 37 0.88-1.00


Agriculture (USDA), total domestic and exportsup ply for U.S. soybeans will grow 73 percentby 1985.

Soybeans are primar ily consumed as animalfeed. But while only 4 percent of their annualprod uction are d irectly consumed by hu mans,the market is growing significantly. The in-troduction of improved textured soy protein incereals, in m eat substitutes and extenders, andin dairy su bstitutes has increased the u se of soyproducts. Nevertheless, the market does not de-

mand soy products in particular but proteinsupplements, vegetable oils, feed grain supple-ments, and meat extenders in general. Otherprotein and oil sources could replace soybeansif the econom ics w ere attractive enough . Fish-meal, dry beans, SCP, and cereals are all poten-tial comp etitors. As long as a substitute canmeet the nutritional, flavor, toxicity, and regula-tory standards, competition will be primarilybased on price.

The comp etition betw een soybeans and SCPillustrates one of the paradoxes of genetic engi-neering. While significant research is attemp t-ing the genetic improvement of soybeans, ge-netic techniqu es are also being explored to in-crease the production of SCP. Consequently, thesame tool—genetic engineering—encouragescompetition between the two commod ities.



Ch. 6—The Food Processing Industry q 109

Gene t i c e ng ine er ing an d

SC P produ c t ion

Despite the m icrobial screening stu dies thathave been conducted and the wealth of basicgenetic knowledge available about common

yeast (a major source of SCP), genetic engineer-ing has had little economic impact on SCP proc-esses until recently. Today, a variety of sub-stances are being considered as raw materialsfor conversion.

q Petroleum-based hydrocarbons.—Until r e -cently, the wide availability and low cost of petrochemicals have mad e the n-alkane hy-drocarbons (straight chain molecules of carbon and hydrogen), which are petro-chemical byprod ucts, potential raw materi-als for SCP production. At British Petro-leum, mutants of micro-organisms havebeen obtained having an increased proteincontent. Mutants have also been foundwith other increased nu tritive values, e.g.,vitamin content.

q Methane or methanol-Relatively few ge-netic studies have been directed at investi-gating th e genetic control of the m icrobialuse of methane or methanol. However, onerecent application of genetic engineeringhas been reported by the Imperia] Chem-ical Indu stries (ICI) in the Un ited Kingdom,where the genetic makeup of a bacterium

(Methylophilus methylotrophus) has beenaltered so that the organism can growmore readily on methanol. The increase ingrowth provides increased protein and hasmade its production less expensive. Thegenetic alteration was accomplished bytransferring a gene from Escherichia coli to

M. methylotrophus.q Carbohydrates. —Many carbohydrate su b-

strates—from starch an d cellulose to beetsand papermill wastes—have been investi-gated. Forests are the most abundantsource of carbohyd rate in the form of cel-

lulose. But before it can be u sed by m icro-organisms, it must be transformed into thecarbohydr ate, glucose, by chemical or en-zymatic pretreatment. Many of the SCPprocesses that use cellulose emp loy orga-

nisms that produce the enzyme cellulase,which degrades cellulose to glucose.

Most of the significant genetic studies on theproduction of cellulase by micro-organisms are just beginning to appear in the literature. Themost recent experiments have been successfulin creating fungal mutants that p rodu ce excessamounts.

C omme rc ia l p roduc t ion

Of the estimated 2 million tons of SCP pro-duced annually throughout the world, mostcomes from cane and beet molasses, with about500,000 tons from hydrolyzed wood wastes,corn trash, and papermill wastes. (See table 23.)

Integrated systems can be designed to couplethe production of a product or food with SCP

production from wastes. E.g., the waste saw-du st from the lum ber industry could become asource of cellulose for micro-organisms. ICI’ssuccessful genetic engineering of a micro-orga-nism to increase the usefulness of one rawmaterial (methanol) should encourage similarattempts for other raw materials.

But wh ile SCP can be obtained from a w idevariety of micro-organisms and raw m aterials,the nutritional value and the safety of eachmicro-organism vary w idely, as do the costs of competing protein sources in regional markets.Consequently, accurate predictions cannot bemad e about the likelihood tha t SCP w ill displacetrad itional protein p rodu cts, overall. Displace-ments h ave and will continue to occur on a case-by-case basis.

Table 23.-Raw Materials Already Tested on aLaboratory or Small Plant Scale

Agave juicesBarley strawCassavaCitrus wastes

Date carbohydratesMeatpacking wastesMesquite woodPeat (treated)

Pulpmill wastesSawdustSunflower seed husks

(treated)

Wastes from chemicalproduction of maleicanhydride

Waste polyethylene (treated)





G e n e t i c s i n b a k i n g , b r e w i n g , a n d w i n e m a k i n g

The micro-organism of greatest significancein the baking, brewing, and winemaking indus-tries is common yeast. Because of its impor-

tance, yeast was one of the first micro-orga-nisms to be u sed in genetic research. Neverthe-less, the surge in studies in yeast genetics hasnot been accompanied by an increase in itspractical application, for three reasons:

q

q

q

industries already have the desired effi-cient strains, mainly as a result of trial-and-error studies;new genetic strains are not easily bred;they are incompatible for mating and thegenetic characteristics are poorly under-stood; andmany of the important characteristics of

industrial microbes are complex; severalgenes being responsible for each.

Changing technologies in the brewing indus-try and increased sophistication in the molec-ular genetics of yeast have m ade it p ossible forresearchers to achieve novel goals in yeastbreeding. one strain that h as already been con-structed can produce a low-carbohydraate beersuitable for diabetics. (See figure 26. )

The baking industry is also und ergoing tech-nological revolution, and yeasts with new prop -erties are now needed for the faster fermenta-tion of dough . New strains with improved bio-logical activity, storage stability, and yieldwould allow improvements in the baking proc-ess.

In the past, most genetic applications havecome in the formation of hybrid yeasts. Thenewel* genetic approaches, which use cell fu-sion now open up the possibility of hybrid s de-veloped from strains of yeast that carry useful

genes but cannot mate normally,

Classical genetic research has also been car-

Figure 26.—The Use of Hybridization To Obtaina Yeast Strain for the Production of

Low-Carbohydrate Beer


ried ou t w ith wine yeasts. Interestingly, withinthe past 10 years, scientists have isolated in-du ced mu tants of wine yeasts that have: I) anincreased alcohol tolerance and the capacity tocompletely ferment grape extracts of unusuallyhigh sugar content; 2) impr oved sed imentation

properties, improving or facilitating separationof yeasts from the wine; and 3) improved per-formance in the production of certain types of wines. Hybridization studies of wine yeastshave been actively pur sued only recently.

Progress in developing strains of yeast w ithnovel properties is limited by the lack of enoughsuitable approved systems for using recombi-nant DN A (rDNA) technology. Eventual ap prov-al by the Recombinant DNA Advisory Commit-tee is expected to boost applied research for thebrewing, baking, and w inemaking industries.




dustry, particularly for starch processing. En-zymes began to be used in quantity only 20years ago. In the early 1960’s, glucoamylase en-zyme treatment began to replace traditionalacid treatment in processing starch; around1965, a stable protease (an enzyme) was in-

troduced into detergent preparations to helpbreak down certain stains; and in the 1970’s,glucose isomerase was u sed to convert glucoseto fructose, practically creating the high-fruc-tose corn syrup industry.

Gene t i c e ng ine er ing a nd e nzy me s in

the food proc e ss ing indus t ry

Biotechnology applied to fermentation proc-esses will make available larger quantities of ex-isting enzymes as well as new ones. (See ch. 5.)The role of genetic engineering in opening com-

mercial possibilities in the food processing in-du stry is illustrated by the enzym e, pullulanase.This enzyme degrades pullulan, a polysaccha-ride, to the ma ltose or high-maltose syrups thatgive jams and jellies improved color and bril-liance. They reduce off-color development pro-duced by heat in candies and prevent sandinessin ice cream by inhibiting sugar crystallization.Mahose has several unique and favorable char-acteristics. It is the least water-absorbent of themaltose sugars and , althou gh it is not as sweetas glucose, it has a more acceptable taste. It isalso fermen table, nonviscous, and easily solu-

ble. It does not readily crystallize and gives de-sirable browning reactions.

Pullulanase can also break dow n an other car-bohydrate, amylopectin, to produce high amy -lose starches. These starches are used in ind us-try as quick-setting, structurally stable gels, asbinders for strong transparent films, and ascoatings. Their acetate derivatives are added totextile finishes, sizing, adhesives, and binders.In food, amylose starches thicken and give tex-ture to gumdrop candies and sauces, reduce fatand grease in fried foods, and stabilize the pro-

tein, nutrients, colors, and flavors in reconsti-tuted products like meat analogs.

In view of the current shortages of petro-leum-derived plastics and the need for a biode-gradable replacement, amylose’s ability to form

plastic-like wraps may provide its largest indus-trial market, although that m arket has not yetbeen developed.

[f applications for the products made bypu llulanase can be d eveloped, genetic engineer-ing can be used to insert this enzyme into in-

dustrially useful organisms and to increase itsproduction. However, the food processing in-

dustry is permitted to use only enzymes that areobtained from sources approved for food use.Since the chief source of pullulanase is a patho-genic bacteriu m, KlebsiellaII aerogenes, no signifi-cant efforts have been mad e to ap ply genetics toimprove its production or quauality. Moleculargenetics could ultimately transfer the pullula-nase trait from K. aerogenes to a micro-organismapproved for food use, if approved micro-orga-nisms that m anufacture p ullulanase cannot befound.

Sweeteners , f lavors , and fragrances

Biotechnology has already had a marked im-pact on the sw eetener indu stry. The availabilityof the enzymes glucose isomerase, invertase,and amylase has mad e the produ ction of high-fructose corn sweeteners (HFCS) profitable. Pro-

duction of HFCS in the United States has in-creased from virtually nothing in 1970 to 10percent of the entire production of caloricsweeteners in 1980 (11 lb per cap ita). The p riceadvantage of HFCS is expected to cause its con-

tinued growth, particularly in the beverage in-dustry. In fact, the Coca Cola Co. announced in1980 that fructose will soon constitute as m uchas 50 percent of the sweetener used in its namebrand beverage.

Biotechnology can be used to pr odu ce othersweeteners as w ell. While it is un likely that su -crose will ever be made by micro-organisms (al-though imp rovements in sugarcane and sugarbeet yields may result from agricultural geneticstudies, see ch. 8.), the microbial production of low-caloric sweeteners is a distinct possibility.

Three new experimental sweeteners—aspar-tame, monellin, and thaum atin—are candidates.

Aspartame is synthesized chemically fromthe amino acids, aspartic acid and phenylala-nine, which can themselves be made by fermen-



114 q Impacts of Applied Genetics-Micro-Organisms, Plants, and Animals

You don ’t worry abou t processing bacon Genetic engineering can be expected to aid inwithou t nitrites, you engineer a synthetic bacon the creation of novel food preparations throughwith designed-in shelf life. effects on both th e food itself and the ad ditives

You d on’t try to educate people to eat a “bal- used for texturizing, flavoring, and preserving.anced d iet;” you create a “whole” food with theproper balance of nutrients and supplements,and you m ake it taste like something p eople

already like to eat.



Ch a p t e r 7

T h e U s e of Gene t i ca l ly

E n gin e e r e d Mic r o -O r g a n is m s

i n t h e En v i r o nm e n t



C h a p t e r 7

Page

Mineral Leach in g an d R ecovery . . . . . . . . . . , . . 117Microbial Leaching . . . . . . . . . . . . . . . . . . . . . 117App lied Gen etics in Strain Imp rovement . . , . . 118Metal Recovery . . . . , . . . . . . . . . . . . . . . . . . . 118

Oil Recovery . . . . . . . . . . . . . . . . . . . . . . . . . .. .119

Enhanced Oil Recovery . . . . . . . . . . . . . . . . . . 119Microbial Production of Chemicals Used

in EOR . . . . , . . . . . . . . . . . . . , . . . . . , . , , 120In Situ Use of Micro-Organisms. . . , . . . . . . . 121EOR and Genetic Engineering . ...........122Constraints to Applying Genetic

Engineering Technologies in EOR. . ......122Genetic Engineering of Micro-Organisms

for Use in other Aspects of Oil Recoveryand Treatment . . . . . . . . . . . . . . . . . . . . . . 123

Overview of Genetic Engineering in Miningand Oil Recovery . . . . . . . . . . . . . , . . . . . . 123

Page

Pollution Control. . . . . . . . . . . . . . . . . . . . . . . . . 123Enhancing Existing Microbial Degradation

Activity. . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1.24Add ing M icrobes to Clean Up Pollu tion . . . . . . 124

Commercial Applications—Market Size and

Prospects . , . . . . . , . . . . . . . . . . . . . . . . . . . 125Genetic Research in Pollution Control . . . . . . . 126Federal Research Supp ort for Engineering

Microbes to Detoxify Hazardous Substances . 127Summary. . . . . . . , . . . . , . . . . . . . . . . . . . . . . 127

Issue and Options—Biotechnology, . . . . . . . . . . . 128

Figure

Figure No. Page

27. chemical Flooding Process . . . . . . . . . . . . . . 120



Ch a p t e r 7

The U s e o f G ene t i ca l l y Eng i nee red

M i c r o - O r g a n i s m s i n t h e E n v i r o n m e n t

Although most gen etically engineered m icro- q

organisms are being designed for contained fa-cilities like fermenters, some are being exam- q

ined for their usefulness in the open environ- q

ment for such pu rposes as mineral leaching andrecovery, oil recovery, and pollution control.

All three app lications are characterized by:

q the use of large volumes of micro-orga-nisms;

less control over the behavior and fate of the micro-organisms;

a possibility of ecological disruption; andless basic research and development (R&D)—and a higher d egree of speculation—thanthe indu stries previously discussed.

M i n e r a l l e a c h i n g a n d r e c o v e r y

All micro-organisms interact with metals,Two interactions that are of potential economicand industrial interest are leaching metals fromtheir ores, and concentrating metals fromwastes or dilute m ixtures. The first w ould allowthe extraction of metals from large quantities of low-grade ores; the second w ould p rovide meth-ods for recycling precious metals and control-ling pollution caused by toxic metals.

Microbia l leaching

In microbial or bacterial leaching, metals inores are m ade soluble by bacterial action. Evenbefore bacterial leaching systems became ac-cepted industrial practice, it was known thatdissolved metals could be recovered from mineand coal wastes. Active mining operations cur-rently based on this process (such as those inRio Tinto, Spain) date back to the 18th century.Present ly, large-scale oper ations in the United

States u se bacterial leaching to recover copp er“rem w aste material. Estimates for the contri-

bution of copper leaching to the total annualJ.S. production range from 11.5 to 15 percent.

Leaching begins with the circulation of waterthrough large quantities–often hundreds of ins—of ore. Bacteria, which are naturally asso-iated with the rocks, then cause the metals to

be leached by one of two general m echanism s:either the bacteria act directly on the ore to ex-tract the metal or they produce substances,such as ferric iron and sulfuric acid, which thenextract the metal. It appears that simply addingacid is not as efficient as using live bacteria.Although acid certainly plays a role in metal ex-traction, it is possible that direct bacterial attack on some ores is also involved. In fact, some of the bacteria that are known to be involved inmineral leaching have been show n to bind tena-ciously to those minerals.

The application of the leaching process touranium mining is of particular interest be-cause of the p ossibility of in situ mining . Insteadof using conventional techniques to haul urani-um ore to the surface, microbial suspensionscan extract the metal from its geological setting.Water is percolated through undergroundshafts where the bacteria dissolve the metals.The solution is then pumped to the surfacewhere the metal is recovered. This approach,also called “underground solution mining, ” is

already used in Canadian uranium m i n e s ,where it began alm ost by chan ce. In 1960, afteronly 2 years of operation, researchers at theStanrock Uranium Mine found that the naturalund erground water contained large amounts of leached uranium. In 1962, over 13,000 kilo-

117

76-565 0 - 81 - 9




grams (kg) of uranium oxide were obtainedfrom the water. Thereafter, water was circu-lated through the mines as part of the miningoperation. It has been suggested that extendingthis practice to most m ines wou ld have signifi-cant environmental benefits because of the

minimal d isrupt ion of the land su r face .Although the process is slower than the technol-ogy currently employed, the operating costsmight be lower because of the simp licity of thesystem, since no grinding machinery is needed.Furthermore, deeper and lower grade d epositscould be mined more read ily.

Bacterial leaching can also extract sulfur-con-taining compounds, such as pyrite, from coal,producing coal with a lower sulfur content.Sulfur-containing coals from such areas as Ohioand the Appalachian Mountains are now less de-

sirable than other coals because of the sulfurdioxide they release du ring burn ing. They oftencontain up to 6 percent sulfur, of which 70 per-cent can be in the form of pyrite. According torecent data, mixed populations of different bac-teria, rather than a single species, are respon-sible for the most effective removal of sulfur—afind ing that m ay lead to th e genetic engineeringof a single sulfur-removing bacterium in thefuture.

App l i e d ge ne t i cs in s t ra in im prov em e n t

The bacterium most studied for its leachingproperties has been Thiobacillus ferrooxidians(which leaches copper), but others have alsobeen identified in natural leaching systems.Although leaching ability is probably undergenetic control in these organisms, practicallynothing is known about the precise mecha-nisms. This is largely because little informationexists in two critical areas: the chemistry of in-teraction between the bacteria and rock sur-faces; and the genetic structure of the micro-organisms. The finding that mixed populationsof bacteria interact to increase leaching efficien-

cy complicates the investigation.Because of the lack of genetic and biochemi-

cal information about these bacteria, the appli-cation of genetic technologies to mineral leach-ing remains speculative. Progress in obtaining

more information is slow because less than adozen laboratories in the Nation are activelyperforming research.

But even when the scientific knowledge isgathered, two obstacles to the use of geneticallyengineered micro-organisms will remain. Thefirst is the need to d evelop engineered systemson a scale large enough to exploit their biologi-cal activities. A constant interchange m ust takeplace between microbial geneticists, geologists,chemists, and engineers. E.g., the geneticistsmust understand the needs identified by thegeologists as well as the p roblems faced by theengineers, who mu st scale-up laboratory-scaleprocesses. The complex natu re of the p roblemcan be approached most successfully by aninterdisciplinary group that recognizes theneeds and limitations of each discipline.

The second obstacle is environmental. In-troducing large numbers of genetically engi-neered micro-organisms into the environmentraises questions of possible ecological disrup-tion, and liability if dam age occurs to the envi-ronment or hum an health.

In summary, the present lack of sufficientscientific knowledge, scientists, and interdis-ciplinary teams, and the concerns for ecologicalsafety p resent the m ajor obstacles to the u se of genetic engineering in microbial leaching.

Metal recovery

The use of micro-organisms to concentratemetals from dilute solutions such as individualwaste stream s has tw o goals: to recover metalsas part of a recycling p rocess; and to eliminateany m etal that may be a pollutan t. The processmakes u se of the ability of micro-organisms tobind metals to their surfaces and then concen-trate them internally.

Stud ies at the Oak Ridge N ational Laboratoryin Tennessee have shown that micro-organismscan be used to remove heavy metals from ind us-trial effluents. Metals such as cobalt, nickel,silver, gold, uranium, and plutonium in concentrations of less than 1 par t per m illion (pp m) car.be recovered. The process is particularly usefulfor recovering metals from d ilute solutions 01



Ch. 7—The Use of Genetically Engineered Micro-Organisms in the Environment q 119

10 to 100 ppm, where nonbiological methods As with other biological systems, genetic

may be uneconomical. Organisms such as the engineering may increase the efficiency of the

common yeast Saccharomyces cerevisiae can ac- extraction process. In the Saccharomyces sys-

cumulate uranium up to 20 percent of their tern, differences in the ability to recover the

total weight. metals have been demonstrated within popula- —

tions of cells. Selection for cells with the genetic

The economic competitiveness of biological ability to accumulate large amounts of specific,methods h as not yet been proven, but genetic desired metals would be an important step inimprovements have been attempted only re- designing a practical system.cently. The cost of producing the micro-orga-nisms h as been a major consideration. If it canbe reduced, however, theuseful.

Oi l r ecovery

app roach might be

Since 1970, oil production in the United

States has d eclined steadily. The supp ly can beincreased by: accelerating explorations for newoilfields; by m ining oil shale and coal and con-verting them to liquids; and by d eveloping newmethod s for recovering oil from existing reser-voirs.

In primary methods of oil recovery, naturalexpulsive forces (such as physical expansion)dr ive the oil out of the formation. In second arymethods of recovery, a fluid such as water ornatural gas is injected into the reservoir to forcethe oil to the well. Approximately 50 percent of

domestic crude in recent years has been ob-tained through secondary recovery.

Recently, new method s of oil recovery havebeen added to primary and secondary m ethods,which are called tertiary, improved, or en-hanced oil recovery (EOR) techniques. They em-ploy chemical and physical methods that in-crease the mobility of oil, making it easier forother forces to drive it out of the ground. Themajor target for EOR is the oil found in sand-stone and limestone formations. It is here thatapplied genetics may p lay a major role ,engineering micro-organisms to aid in recovery.

Oil susceptible to these processes is localizedin reservoirs and pools at depths ranging from100 ft to mor e than 17,000 ft. In these areas, theoil is adsorbed on grains of rock, almost alwaysaccompanied by water and natural gas. The

physical association of the trapped oil and the

surrounding geological formations varies signif-icantly from site to site. The unknown charac-teristics of these variations ar e largely respon-sible for the economic risk in an attempted EOR.

Enhanced o i l recovery

Of the original estimated volume of morethan 450 billion barrels (bbl) of U.S. oil reserves,about 120 billion bbl have been recovered byprimary and secondary techniques, and another30 billion bbl ar e still accessible by thesemethods. The remaining 300 billion bbl how-

ever, are probably recoverable only by EORmethod s. These figures include the oil remain-ing in known sandstone and limestone reser-voirs and exclud e tar sand s and oil shale.

Four EOR processes are currently used. Allar e designed to dislodge the crud e oil from it s

natural geological setting:

q

q

In thermal processes, the oil reservoir is

heated, wh ich causes the v iscosity of the oi l

to decrease, and with the a id of thepressure of the air introduced, supportsthe combustion that forces the petroleum

to the producing well. Thermal processeswill not be improved by genetic technol-ogies.Various crud e oils differ in their viscosity—ability to flow. Primary and secondarymethods can easily remove those that flow



Ch. 7—The Use of Genetically Engineered Micro-Organisms in the Environment . 121

obtained by microbial fermentation could com-pete with those obtained from alternativesources, especially seaweed. Controlled fermen-tation is not affected by marine pollution andweather, and production could be geared tomarket demand.

Biological p rocesses have disadvantages pri-marily in the costs of app ropriate raw materialsand in the need for large quantities of solvent.Current efforts to find cheaper raw materials,such as sugar beet pulp and starch, show prom-ise. The need for solvents to precipitate and con-centrate the polymers before shipment fromplant to field can be circumven ted by p rodu cingthem onsite.

Micro-organisms can also produce substanceslike butyl and p ropyl alcohols that can be u sedas cosurfactants in EOR. It has been calculatedthat if n-butanol were used to prod uce crude oilat a level of 5 percent of U.S. consumption, 2billion to 4 billion lb per year—or four to eighttimes the current butanol production—wouldbe required. Micro-organisms capable of pro-ducing such surfactants have been identified,and genetically superior strains were isolatedseveral decades ago at the Northern RegionalResearch Laboratories in Illinois. Other chem-icals, such as alcohols that increase the rate of formation and stability of chemical/ crude oilmixtures and the agents that help prevent pre-cipitation of the surfactants, have also been pro-duced by microbial systems.

The uncertainties of the technical and eco-nomic parameters are compounded by the lack of sufficient field experiments. Laboratory testscannot be equated w ith conditions in actual oilwells. Each oil field has its own set of character-istics—salinity, pH (acidity and alkalinity),temperature, porosity of the rock, and of thecrude oil itself—and an injected chemical be-haves differently in each setting. In most cases,not enough is known about a well’s characteris-tics to predict the nature of the chemical/ crude

oil interaction and to forecast the efficiency of oil recovery.

use w ater available at site; growth u nder cond itions that discour-age the growth of un wanted m icro-organisms; no m ajor problemsin culturing the bacterium; and genetic stability.

IN SITU USE OF MICRO-ORGANISMS

One alternative to growing m icro-organismsin large fermenters then extracting their chem-ical products and injecting them into wells, is toinject the micro-organisms directly into thewells. They could then produce their chemicals

in situ.

Unfortunately, the geophysical and geochem-ical conditions in a reservoir seldom favor thegrowth of micro-organisms. High temperature,the presence of sulfur and salt, low oxygen andwater, extremes of pH, and significant engi-neering hurdles make it difficult to overcomethese limitations. The micro-organisms must befed and the microenvironment must be care-fully ad justed to their needs a t d istances of hun-dred s to thousan ds of feet. The oil industry hasalready had discouraging experiences with

micro-organisms in the past. In the late 1940’s,for instance, the injection of sulfite-reducingmicro-organisms, along with an inadvertentlyhigh-iron molasses as a carbon source, resultedin the formation of iron su lfide, wh ich cloggedthe rock pores. One oil company developed ayeast to break dow n p etroleum, but the size of the yeast cells (5 to 10 micrometers, um) wasenough to clog the 1-um pores.

Neverth eless, information from geom icrobi-ology suggests that this approach is worth pur-suing. Preliminary field tests have also been en-couraging. The injection of 1 to 10 gal of Bacillusor Clostridium species, along with a water-suspend ed mixture of fermentable raw materi-als such as cattle feed molasses and mineralnutrients, has resulted in copious amounts of carbon dioxide, methane, and some nitrogen inreservoirs. The carbon d ioxide m ade the crud eless viscous, and the other gases helped torepressurize the reservoir. In addition, largeamou nts of organic acids formed add itional car-bon dioxide through reactions with carbonateminerals. The production of microbial sur-factants further aid ed the p rocess.

Although previous assessments have arguedthat reservoir pressure is a significant hin-drance to the growth of micro-organisms, morerecent studies indicate the contrary. The micro-organisms must, however, be selected for in-creased salt and pH tolerance.




EOR AN D GENETIC ENGINEERING

The current research approach, funded bythe Departm ent of Energy (DOE) and, independ -ently, by various oil comp anies, is a tw o-phaseprocess. The first phase is to find a micro-organism that can function in an oil reservoir

environment with as many of the necessarycharacteristics as possible. The second is to alterit genetically to enhan ce its overall capability.

The genetic alteration of micro-organisms toprod uce chemicals used in EOR has been mor esuccessful than the alteration of those that maybe used in situ. * However, recombinant DNA

(rDNA) technology has not been ap plied in ei-ther category. All efforts have em ployed artifi-cially indu ced or n aturally occurring m utations.

CONSTRAINTS TO APPLYING GENETICENGINEERING TECHN OLOGIES IN EOR

The genetic data base for micro-organismsthat produce useful polysaccharides is weak.Few genetic stud ies have been d one. Hence, the-oretically plausible approaches such as transfer-ring enzyme-coding plasmids (see ch. 2) forpolysaccharide synthesis, cannot be seriouslycontemplated at present. Only the crudestmethods of genetic selection for desirable prop-erties have been used thus far. They remain theonly avenue for improvement until more islearned about the micro-organism’s geneticmechanisms.

The biochemical data base for the character-istics of both the micro-organisms and theirprod ucts is also lacking. The wide potent ial forchemical reactions carried out by m icrobes re-mains to be explored. At the sam e time, a sys-tem m ust be d evised to allow easy characteriza-tion, classification, and comparison of productsderived from a variety of micro-organisms.

The physical data base for oil reservoirs islimited. The u niqueness of each reservoir sug-gests that no u niversal micro-organism or meth-od of oil recovery will be found. Compounding

q Some of the goals have b een to: improve polymer properties toenhan ce their comm ercial applicability; improve polymer produc-tion (a major mistake h as been to reject a m icro-organism in theinitial screening b ecause its level of production was too low ); im-prove culture characteristics, e.g., resistance to phage, rapidgrowth, ability to use cheaper raw materials; and eliminate en -zymes that naturally degrade the polymers.

this problem is the lack of sufficient physical,chemical, and biological information abou t thereservoirs, without which it is difficult to seehow a rational genetic scheme can be con-structed for strains. Clearly, the activities of micro-organisms under specified field condi-

tions cannot be studied unless researchersknow what the appropriate conditions are.

Three institutional obstacles exist. First, publi-cation in this field is limited because most re-search is carried out in the commercial worldand remains largely confidential. Second, nei-ther the private nor the public sector has beenenthusiastic about the potential role of micro-organisms in EOR, The biological approach hasonly recently been given consideration as a w ayto advance the state of the art of the technology,and most oil compan ies still have limited staffs

in microbiology. To date, DOE’s Division of Fossil Fuel Extraction has condu cted the m ainFederal effort. Third, any effort to use micro-organisms must be multidisciplinary in nature.Geologists, microbiologists (including microbialphysiologists and geneticists), chemists, andengineers must interact to evolve successfulschemes of oil recovery. Thus far, such teamsdo n ot exist.

Environmental and legal concerns have also in-hibited progress. Microbial EOR methods usual-ly require significant quantities of fresh water

and thus may compete with municipal and agri-cultural uses. Furthermore, the use of micro-organisms introduces concerns for safety. Allstrains of Xanthomonas, which produce xanthangum polymer , are plant pathogens. Othermicro-organisms with potential, such as Scleroti-um rolfii and various species of Aureobasidiumhave been associated with lung disease andwound infections, respectively.

Immed iate environmental and legal concerns,therefore, arise from the p otential risks associ-ated with the release of micro-organisms intothe environment. When they naturally cause

disease or environmental d isruption, their use isclearly limited. And w hen they do not, geneticengineering raises the possibility that theymight. Such concerns have reduced th e privatesector’s enthusiasm for attempting genetic




engineering. (See ch. 10 for a more detaileddiscussion of risk.)

GENETIC ENGINEERING OF MICRO-ORGANISMSFOR USE IN OTH ER ASPECTS OF OIL

RECOVERY AND TREATMENT

Two other aspects of microbial physiology

deserve attention: the microbial production of oil muds or drill lubricants, and the treatmentof oil once it has been recovered. Drilling mudsare suspensions of clays and other materialsthat serve both to lubricate the drill and tocounterbalance the upward pressure of oil. Mi-crobially produced polysaccharides have beendeveloped for this use. Exxon holds a p atent ona formulation based on the production of xan-than gum, from Xanthomonas campestris, whilethe Pillsbury Co. has developed a polysac-charide (glucan) from various species of Scler- otium. At least two of the smal l gene t icengineering firms have begun research pro-grams to develop biologically p rodu ced p olysac-charides with the d esired lubricant q ualities.

Interest in the postrecovery microbial treat-ment of oil after its extraction centers aroundthe ability of micro-organisms to remove un-

desirable constituen ts from the crud e oil itself.As an indication of recent progress, three dis-tinct microbial systems have been developed tohelp remove aromatic sulfur-containing mate-rial, a major impurity.

Overv iew of gene t ic engineer ing inmin ing and o i l r e c ov e ry

The underlying technical problem with theuse of genetically engineered organisms ineither mining or oil recovery is the magnitudeof the effort. In both cases, large areas of landand large volumes of materials (chemicals, flu-ids, micro-organisms) must be used. The resultsof testing any new micro-organism in a labora-tory cannot automatically be extrapolated tolarge-scale applications . T h e c h a n g e i nmagnitude is further complicated by the lack of

rigid controls. Unlike a large fermen ter wh osetemperature, pH, and other characteristics canbe carefully regulated, the natural environmentcannot be controlled. Nevertheless, despite theformidable obstacles, the poten tial value of theproducts in these areas assures continuing ef-forts.

P o l l u t i o n c o n t r o l

Life is a cycle of synthesis and degradation—

synthesis of complex molecules from atoms andsimple molecules and degradation by bacteria

yeast, and fungi, back to simpler molecules and

atoms when organisms die. The degradation of

complex molecules is an essential part of life.

Without it, “. . . we’d be knee-deep in dino-

saurs. ”1 A more q uantitative statement is equ al-

ly thought provoking. Livestock in the united

States produce 1.7 billion tons of manure an-

nu ally. Almost all of it is degrad ed b y soil micro-

organisms.

For a long time people have exploited micro-

bial life forms to degrade and detoxify humansewage. Now, on a smaller scale, science is

‘R. B. Grubbs, “Bacterial Supplementation, What It Can and Can-

not Do. ” oral presentation to the Ninth Engineering Foundation on

Environmental Engineering in the Food Processing Industry, 1979(Available from Flow Laboratories, Inc., Rockville, Md.)

beginning to u se micro-organisms to deal with

the pollution problems p resented by indu strialtoxic wastes. Chemicals in their place can beuseful and beneficial; out of place, they can bepolluting.

Pollution problems can be divided into twocategories: those that have been present for along time in the biosphere-e. g., most hydro-carbons encountered in the petroleum industryand human and animal wastes—and those thatowe their origin to human inventiveness-e. g.,certain pesticides. Chemicals of both sorts,through mishap, poor planning, or lack of

knowledge at the time of their applicationsometimes appear in places where they arepotentially or actually hazardous to humanhealth or the environment.

Pollution can be controlled by microbes intwo w ays: by enhancing the growth and activity




pounds of bacteria is unlikely to have any effect.Thus far, the Environmental Protection Agency(EPA) has not recommended adding bacteria tomunicipal systems; however, EPA suggests thatthey might be useful in smaller installations andfor specific problems in large systems.

Dry formulations are available for use incleaning drains and pipes in smaller installa-tions, such as restaurants and other food proc-essing facilities. In restaurants, the bacteria areadd ed to the drain at the end of the workday.Bacteria have been selected for their inability toproduce hydrogen sulfide, which means thatthe degrading process does not produce the un-pleasant odors frequently encountered in thedigestion of oils and fats.

6

As of November 1979, the pollution controlindustry had few plans for the genetic manipu-

lation of bacteria, except for the selection of naturally occurring better performers. Con-sumer resistance to “mutants” is a factor thatdiscourages the move to microbial genetics.Probably even more important is the high costof establishing and maintaining microbial genet-ics laboratories. It has been estimated that thecost of carrying a single Ph. D. microbial geneti-cist is over $100,000 annually.

7This expense is

qu ite high r elative to the $2 million to $4 millionsales of all biological pollution control com-panies in 1978.

8

Resistance to the use of genetically manip-ulated bacteria is not un iversal. Many ind ustrialwastes are oxidized to nontoxic chemicals bybiological treatment in aerated lagoons. Theprocess depends on the presence of microbes inthe lagoons; over time, those that grow best onthe w astes come to d ominate the m icrobial pop -ulations. Three companies now sell bacteriathat they claim outperform the indigenousstrains found in the lagoons. E.g., the Polybac

Corp. has sold its products to all seven Exxonbiological waste treatment plants to treat chem-ical wastes. One of its formulations has beenused to degrad e toxic dioxins from an herbicidespill. One m onth ’s treatment w ith the bacterialformulation reduced the orthochlorophenol

concentration from 600 to 25 ppm in a 20,000 -gal lagoon.

9

Sybron/ Biochemical, a division of SybronCorp., sells cultures of bacteria that are in-tended to aid in the biological oxidation of in-du strial wastewater; this comp any also lists 20different cultures for application to specificwastes. Patent number 4,199,444 was grantedon April 22, 1980, for a process involving theuse of a mutant bacterial culture to decolorwaste water p rodu ced in Kraft p aper p rocess-ing.

l0Other patents are pending on a m ixture of

two strains that degrad e grease and a strain thatdegrades “nonbiodegradable” detergents. 11

There is disagreement about the va lue of add -ing microbes to decontaminate soils or waters.One point of view argues that serious spills fre-quently sterilize soils, and th at ad ding m icrobesis necessary for any biod egradation. The othercontends that encouraging indigenous microbesis more likely to succeed because they are ac-climated to the spill environment. Added bac-teria have a difficult time competing with thealready-present microbial flora. In the case of marine spills, bacteria, yeast, and fungi alreadypresent in the water participate in degradation,no one has been able to demonstrate the useful-ness of add ed m icrobes.

C omme rc ia l app l i c a t ions— mark e t s i z e

and prospe c t s

The estimated market size of pollution-con-trol biological pr odu cts in 1978 was $2 million to

$4 million, divided among some 20 comp anies,

‘Anon., Wean That Sewage System With Bugs!” Environmental

Science and 7’echno/ogy 13:1 198-1199, 1979.7

Anon., “Biotechnology DNA Research Expenditures in U.S. MayReach $500 Million in 1980, With About $150-200 Million for Com-mercial Products, ” Hi l l told. Drug Research Reports, “The BlueShe et, ” M ay 28, 1980, p. 22.

aAnon., Business Week, July 5, 1976, p. 280; Chemical Week 121:47, 1977; an d Food Engineering 49: 138, 1977, cited in T. Gass-ner, “Microorganisms for Waste Treatment, ” Microbial Technol-

~gv, 2 cd., vol. II, (London: Academic Press, 1979), pp. 211-222.

9See footnote 6.IOL. Davi s, J. E. Blair, an d C, W. Randall, “Commu nicant ion:

Development of Color Removal Potential in Organisms TreatingPulp an d Pap er Wastewater,” J. Water Pollution Control Fed., Feb-ru a rv 1978, pp. 382-385.

i IF. Sp rahe r an d N. Tekeocgak , “Foam Control an d Degrada t ionof Nonionic Detergent,” Industrial Wastes, January/February 1980;L. David, J. E. Blair, and C. Randall, ‘(Mixed Bacterial Cu ltures Leak ‘Non-Biodegradable’ Detergent,” industrial Wastes, May/June1979.




and the potential market was estimated to be asmuch as $200 million.

12These estimates can be

compared to Polybac’s own sales records. In1976, its first year, its sales totaled $0.5 millionand in 1977, $1.0 million. It expects to reach $5million in 1981.

To date genetically engineered strains havenot been ap plied to pollution problems. At leastone prominent genetic engineering companyhas decided not to enter the pollution controlfield, concluding that it was improbable thatadded microbes could compete with indigenousorganisms. More specifically, the possibility of liability problems make the approach even lessattractive. Pollution control requires that “new”life forms be released into the environment,which is already seen as precariously balanced.Such new forms might cause health, economic,

or environmental problems. The problems of liability that might arise from such applicationsare enough to deter entrepreneurs from con-templating w ork in the field at this time.

An additional reason for the reluctance of some companies to engage in this activity is thatthe opp ortunities for making m oney are limited.Selling microbes, rather than their products,may well be a one-shot opportunity. The mi-crobes, once purchased, might be propagatedby the buyer. Nevertheless, at least two smallcompanies have announced that they are pursu-ing efforts to use genetic engineering.

The low-key efforts in this field might accel-erate qu ickly if a significant breakthr ough oc-curred, To date, no “new” organism has ap-peared that will degrade previously intractablechemicals. The effect of such a developmentmight be enormous.

Gene t i c re searc h in po l lu t ion c on t r o l

The Oil and Hazardous Materials SpillsBranch of EPA currently supports research

aimed at isolating organisms to degrade threespecific chemical compounds. The work is beingcarried out on contract; as of Novem ber 1979,no field trials of the organisms had been un der-

‘zSee footnote 8.

taken. Two of the toxic chemicals, pentachoro-phenol and hexachlorocyclopentadiene, arerelatively long-lived compounds and presentlong-term problems. A fungus and a bacteriumthat can degrade the first compound have beenisolated,

13and Sybron/ Biochemical already sells

a culture specifically for pentachlorophenoldegrad ation. The third toxic compou nd is meth-yl parathion. Its inclusion is more difficult tounderstand, since it is degraded within a fewdays after its application as a pesticide.

Efforts have been made to isolate bacteriathat can degrade (2,4-dichlorophenoxy) aceticacid (2,4-D) and (2,4,5-trichlorophenoxy) aceticacid (2,4,5-T), the components of AgentOrange.

14Strains of the bacterium Alcaligenes

paradoxus rapidly degrade 2,4-D, and thegenetic information for the degradation activity

has been located on a plasmid. The investigatorwho found that strain, while optimistic aboutthe opp ortun ities for isolating and transferringother resistance genes, has been unable to finda bacterium that degrades 2,4,5-T or its verytoxic contaminant, 2,3,7,8-tetrachlorodibenzo-para-dioxin (TCDD or dioxin).

By far the best kn own research in this area isthat of Dr. Ananda M. Chakrabarty who engi-neered two strains of Pseudomonas, each of wh ich h as the ability to d egrade th e four classesof chemicals found in oil spills. Chakrabartybegan with four different strains of Pseudo-

monas. None of them presented a threat tohum an health, and each could degrade one of the four classes of chemicals. His researchshowed that the genes controlling the d egradingactivities were located on plasmid s. Taking ad -vantage of the relative ease of moving suchgenes among bacteria, he produced two recom-binant bacteria.

Chakrabarty has presented evidence that hisbacterium degrades complex petroleum mix-tures such a s crude oil or “Bunker C“ oil, and he

‘SN. K. Thuma, P. E. O’Nei l l , S. G. Brown lee, and R. S. Valentine,“Laboratory Feasibility and Pilot Plant Studies: Novel Biodegrada-tion Processes for the Ultimate Disposal of Spilled HazardousMaterials, ” National Environment Research Center, U.S. Environ-mental Protection Agency, Cincinnati, Ohio, 1978.

M J. M < pembe r t on , “pesticide Degradin g plasmids : A Biological

Answer to Environmental Pollution byPhenoxyherbicides,” Arnbio

8:202-205, 1979.




has prop osed a method for u sing it to clean upoil spills. The bacteria are to be grown in thelaboratory, mixed with straw, and dried. Thebacteria-coated straw can be stored untilneeded, then dropped from a ship or aircraftonto oil spills. The straw absorbs the oil and the

bacteria degrades it.15

To comp letely cleanup aspill will probably require mechanical efforts inaddition to the biological attack. It was the pro-duction of one of Chakrabarty’s strains that ledto the Sup reme Court d ecision on “ the patentingof life. ” (See ch. 12 for further details.)

The essential difference between the well-publicized Chakrabarty approach and a lesswell-known one is that all the desired activitiesin Chakrabarty’s approach are combined in asingle organism; while in the other method,bacteria bearing single activities are mixed

together to yield a d esired “formulation.” In yetanother approach, Sybron/ Biochemical usesmutation and selection to produce specializeddegradation activities. It also sells mixedcultures for some applications.

The single-organism, multiple-enzyme systemhas the adv antage that every bacterium can at-tack a number of compounds. The mixed for-mulations allow the preferential proliferation of bacteria that feed on the most abu nd ant chem-ical; then, as that chemical is exhausted, otherbacteria, which flourish on the next most abun-dant chemical, become dominant. The pref-

erential survival of only one or a few strains in amixed formulation might result in no bacteriabeing available to degrade some compounds.The multienzyme bacteria, on the other hand,can degrade one chemical after another, oralternatively, more than one at the sam e time.

Federal research support for

engineer ing microbes to de tox i fy

h a z a r d o u s s u b s t a n c e s

EPA currently limits its support to researchaimed at selecting indigenous m icrobes, an area

lspa~ent specification I 436 573, Mav 19, 1976, Patent off ice ,

London, England.

that has already attracted some commercialresearch supp ort. Commercial firms are lookingfor large-scale m arkets, such as sew erage sys-tems, or comm only occurr ing smaller markets,such as gasoline spills and common industrialwastes.

Whatever potential exists in identifying,growing, and using naturally occurring mi-crobes for pollution control pales beside the op-portunities offered by engineering new ones,Unfortunately, the potential risks increase aswell. EPA has taken a preliminary step towar dassessing the risks by soliciting studies to deter-mine w hat env ironmental risks may exist fromaccidentally or deliberately released engineeredmicrobes.

S u m m a r y

While some unreported efforts may beunderway, genetics has apparently been littleapplied to pollution abatement. Nevertheless,the p rodu ction of “new ” life forms that offer asignificant improvement in pollution control is apossibility. The constraints are questions of liability in th e event of health , econom ic, or en-vironmental damage; the contention that addedorganisms ar e not likely to be a significant im-provement; and the assumption that sellingmicrobes rather than products or processes isnot likely to be profitable.

The factors that have discouraged develop-ments in th is area wou ld p robably become lessdeterring if convincing evidence were foundthat microbes could remove or degrade an in-tractable pollutant. In the meantime, the re-search necessary to produce marked improve-ments h as been inhibited. Overcoming th is in-hibition may require a governmental commit-ment to support the research, to buy themicrobes, and to provide for protection againstliability su its. Such a governm ental role wou ld

be in keeping w ith its comm itment to protectinghealth and the environment from the toxic ef-fects of pollutan ts.




I s s u e a n d O p t i o n s — B i o t e c h n o l o g y

I S S U E: H ow c a n the F e de r a l G ove r n -ment promote advances in bio-t e c h n o l o g y a n d g e n e t i c e n g i -n e e r i n g ?

The United States is a leader in ap plying ge-netic engineering and biotechnology to indus-try. One reason is the long-standing commit-ment by the Federal Government to the fundingof basic biological research; several decades of sup port for some of the most esoteric basic re-search has unexpectedly provided the founda-tion for a highly u seful technology. A second isthe availability of venture capital, which has al-lowed th e formation of small innovative compa-nies that can bu ild on the basic research.

The argument for Government p romotion of

biotechnology and genetic engineering is thatFederal help is needed in those high priorityareas not being developed by industry.

The argum ent against such assistance is thatindustry will develop everything of commercialvalue without Federal help.

A look at what indu stry is now attempting in-dicates that sufficient investment capital isavailable to pursu e specific manufacturing objectives, such as for interferon and ethanol, bu tthat some high-risk areas that might be of in-terest to society, such as p ollution control, may

need promotion by the Government. Otherareas, such as continued basic biological re-search, might n ot be profitable soon enou gh toattract industry’s investment, Specialized educa-tion and training are areas in wh ich the Govern-ment has already played a major role, althoughindustry has both supported university trainingand cond ucted its own inhou se training.

OPTIONS:

A. Congress could allocate funds specifically for

genetic engineering and biotechnology R&D in

the budget of appropriate agencies, such as the National Science Foundation (NSF), theU.S. Department of Agriculture (USDA), the

Department of Health and Human Services(DHHS), the Department of Energy (DOE), the

Department of Commerce (DOC), and the De- partment of Defense (DOD).

Congress has a long history of recognizingareas of R&D that need priority treatment inthe allocation of funds. Biotechnology has notbeen one of these. Even though agencies likeNSF receive congressional funding, its Alter-native Biological Sources of Materials programis one of the few applied programs that is notcongressionally mandated. As a result, the fiscalyear 1980 bud get saw a reduction in the alloca-tion of funds, from $4.1 million in 1979 to $2.9million. A congressionally mandated program,analogous to the successful NSF EarthquakeHazard Mitigation program, could be writteninto law. other programs, such as the com-

petitive grants progr am at USDA (or the Officeof Basic Biological Research at DOE), are alsomodestly funded.

Increasing the amount of money in an agen-cy’s biotechnology program could bring criti-cism from other programs within each agency if their levels of funding are not increased com-mensu rately. The Competitive Grants Programat USDA has similar problems; those who aremost critical of it argue that it should not takefunds from traditional programs. Nevertheless,Congress could p romote two types of programs:those w ith long -range payoffs (basic research),and those which industry is not willing to un-dertake but that might be in the national in-terest.

B. Congress could establish a separate Institute of Biotechnology as a funding agency.

The merits of a separate institution lie in thepossibility of coordinating a wide range of ef-forts, all related to biotechnology. Among pres-ent organizations, biotechnology and app lied ge-netics cut across several institutes and divisionswithin them. Med ically oriented research falls

primarily u nd er the dom ain of the National In-stitutes of Health (NIH). EPA is concerned withthe p reven tion of pollution; w hile NSF’s effort inbiotechnology has been restricted to modestsupport scattered through several divisions.




The creation of an organization such as the Na-tional Technology Foundation (H.R. 6910) wouldrepresent the kind of comm itment to engineer-ing, in general, that currently does not exist.

Competition for fun ds w ithin other agencies

wou ld be avoided , since funding w ould now oc-cur at the level of congressional appropriations.A separate institute, carrying the stamp of Government recognition, wou ld make it clear tothe public that this is a major new area withgreat potential. This might foster greater aca-demic and commercial interest in biotechnologyand genetic engineering.

On the other hand , biotechnology and geneticengineering cover such a broad range of disci-plines that a single agency would overlap themandates of existing agencies. Furthermore, thecreation of yet another agency carries with it all

the disadvantages of increased bureaucracy andcompetition for funds at the agency level.

c. Congress could establish research centers inuniversities to foster interdisciplinary ap-

proaches to biotechnology. In addition, a program of training grants could be offered to train scientists in biological engineering.

The successful use of biological techniques in

indust ry depends on a mul t id isc ipl inary ap-

proach involving biochemists, geneticists, mi-

crobiologists, process engineers, and chemists.Little is now being done, publicly or privately,

to develop expertise in this interdisciplinaryarea.

In 1979, President Carter proposed the crea-

tion of generic technology centers (useful to a

broad range of industries) as one way to stim-

ulate innovation. The centers would conduct

the kin d of research that an ind ividual companymight not consider cost effective, but that might

ultimately benefit several companies. Each cen-

ter would be jointly funded by Government andindustry, with Government providing the seed

money and industry carrying most of the costs

within 5 years. If the centers were establishedat universi t ies, startup costs could be mini-

mized.

Several congressional bills contain provisionsfor centers similar to these. For example, on

October 21, 1980, President Carter signed intolaw a bill (S. 1250) that would establish Centersfor Industrial Technology to foster researchlinks between industry and universities. Theywould be affiliated with a university or non-profit institution.

One or more of these centers could be specifi-cally designated to specialize in biotechnology.In addition, training grants could be used tosupp ort the edu cation of biotechnologists at thecenters or elsewhere. Curren tly, there is no na-tionw ide training p rogram to train stud ents inthis discipline. Education programs, especiallyfor the postgraduate and graduate training of engineers, could further the idea of using bio-logical techniques to solve engineering prob-lems.

D. Congress could use tax incentives to stimulate

biotechnology.

The tax laws could be used to stimu late bio-technology in several ways. First, they could ex-pand the supply of capital for small high-risk firms, wh ich are generally considered more in-novative than established firms, because of their willingness to undertake the risks of in-novation. Much of the pioneering work in theindu strial app lication of genetic techn iques hasbeen done by such firms. By nature, they arespeculative, high-risk investments. Second, thetax law could p rovide special subsidies to new

high-technology firms, which cannot use thestandard investment incentives, such as the in-vestment tax credit, because they usually haveno taxable profits for the first several yearsagainst wh ich to app ly the tax credit. Third, taxincentives could be provided for both estab-lished and new firms to make the investment of money for R&D more attractive.

There are a number of ways to expand thesupp ly of venture capital. One is to decrease thetax rate on capital gains or the period an assetmust be held for it to be considered a capitalgain rather than ordinary income. This changecould be limited to stocks in high-technologyfirms in ord er to focus its imp act and m inimizerevenue loss. Other options involving the stock of high-technology compan ies are: a tax creditto the investor wh o pu rchases the stock; defer-




ment of capital gains taxes on th e sales of thesestocks if the p roceeds ar e reinvested into simi-larly qualifying stock; and more liberal capitalloss provisions.

In add ition to focusing on the sup ply of cap-ital, tax policy could attempt to directly increasethe profitability of potential growth companies.Since most are not profitable for several years,they cannot take full advantage of the invest-ment tax credit—or even the p rovision for car-rying net operating losses back 3 years and for-ward 7 years to offset otherwise taxable profits.Two p roposals may remed y this situation. First,the investment tax credit could be r efun dable tothe extent it exceeded any tax liability of thefirm. A preliminary estimate of the revenue lossfor this proposal was $1 billion for 1979. Sec-ond, new companies could be permitted to

carry net operating losses forward for 10 years.This change would give new firms the samenumber of years over which to deduct losses asestablished firms.

The final type of tax incentive is directed atincreasing R&D expend itures. Two major pro-posals would p ermit comp anies to take tax cred-its on a certain percentage of their R&D ex-penses, and on contributions to u niversities forresearch.

The R&D credit has been ad vocated for sev-eral reasons. First, it would increase the after-

tax return on R&D investments, making themmore attractive. Second, it would reduce thedegree of risk on such investments; with a 10-percent credit, the real after-tax expense of a $1million investment is $900,000. Finally, it wouldgive firms m aximum flexibility in selecting p roj-ects for investment.

Questions have been raised about the cost ef-fectiveness of the credit. For calendar year1980, the Treasury Department estimated thecost of a lo-percent R&D credit to be $1.9 mil-lion. Since R&D costs average only 10 to 20 per-

cent of the total cost of bringing a n ew p roductor process to the market, the net reduction inthe cost of commercializing an invention wouldbe 1 to 2 percent. Moreover, the commercialstage of innovation is thou ght to be riskier andcostlier than the technical stage. Another p rob-

lem is that the credit maybe a windfall for firmsthat w ould be investing in R&D anyw ay. Finally,the credit would subsidize R&D devoted tominor product changes or incremental improve-ments in addition to R&D directed to more fun-damental breakthroughs.

One of the provisions of a pending congres-sional bill (H.R. 5829) provid es for a credit of 25percent for incremental research expendituresabove those for a base period. By limiting thecredit to incremental expenditures, the billwould create a more cost-effective credit, if passed.

The final type of tax credit wou ld be for cor-porate contributions to university research. TheTreasury Department estimated that a 25 per-cent credit for research in all fields would cost$40 million in 1980. This credit would be tar-

geted to more fund amental research and not tothe su bsidy of short-term, incremental p rojectsthat are usually a significant part of corporateR&D budgets.

E. Congress could improve the conditions underwhich U.S. companies can collaborate with

academic scientists and make use of the technology developed in universities in whole or in part at the taxpayer expense.

Developments in genetic engineering havekindled interest in this option. Nevertheless, theGovernm ent’s role in fostering un iversity-aca-demic interaction is far from accepted. Such arole may limit the flexibility of a cooperative ef-fort. At the very least, disincentives such as pat-ent restrictions could be removed .

The controversy has been sum med up as fol-lows.

At the next level of involvement, the Govern-ment could ident ify potential partners, and fa-cilitate negotiations. A more active role wouldinvolve the Government’s providing startu pfunds. Finally, the Government could be a th irdpartner, sharing costs with industry and the

university. In this case, too large a Governmentrole could lead to Federal intervention in activ-ities that should be the responsibility of busi-ness and industry.

‘Dennis Prager, G. S. omenn, Science 207: 379-384, 1980,



P a r t II

A g r i c u l t u r e

Chapter 8—The Application of Genetics to Plants . . . . . . . . . . . . . . . . . . . . . . . . . . . , . . . . . . 137

Chapter 9—Advances in Reproductive Biology and Their Effects on Animal Improvement . . . 167



C h a p t e r s

Th e Ap p lic a t ion o f

Ge n e t ic s t o P la n t s



C h a p t e r s Page

Persp ective on Plan t Breed in g. . . . . . . . . . . . . . . 137The Plant Breeder’s Approach to

Commercialization of New Varieties . . . . . . . 138Major Constraints on Crop Imp rovement. . . . . 139

Genetic Technologies as Breeding Tools . ......140New Genetic Technologies for Plant Breeding .141

Phase I: Tissue Culture to Clone Plants ... , . 141

Phase H: Engineering Changes to AlterGenetic Makeup; Selecting Desired Traits. . 144Phase 111: Regenerating Whole Plants From

Cell s in Tissu e Cu ltu re . . . . . . . . . . . . . . . . 146Constraints on the New Genetic Technologies . 149

Technical Constraints. . . . . . . . . . . . . . . . . .149Institu tion al Con straint s . . . . , . . . . . . , . . . . 150

Impacts on Generating New Varieties . . . . . . . . . 151Examples of New Genetic Approaches . ......152Selection of Plants for Metabolic Efficiency .. .152Nitrogen Fixation. . . . . . . . . . . . . . . . . . . . . . . 152

Genetic Variability, Crop Vulnerability, andStorage of Germplasm. . . . . . . . . . . . . . . . . . 154

The Amoun t of Genetic Erosion That Has

Taken Place . . . . . . . . . . . . . . . . . . . . . . . .. 154The Amount of Germplasm Needed . ........154The National Germplasm System . . . . . . . . . . . 155The Basis for G eneti c Uniform ity . . . . . . . . . . . 157Six Factors Affecting Adequate Management

of Genetic Resources. . . . . . . . . . . . . . . . . . . 158

Page

Summary . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 160Issues and Options—Plants . . . . . . . . . . . . . . 161Technical Notes . . . . . . . . . . . . . . . . . . . . . 162

TablesTable No. Page

24. Average Yield per Acre of Major Crops in

1930 and 1975. , . . . . . . . . . . . . . . . . . . . . . 1372.5. Some Plants Propagated Through TissueCultu re for Prod uction or Breedin g. . . . . . . 143

26. Representative List of Tissue CulturePrograms of Commercial Significance in

the United States. . . . . . . , . . . . . . . . . . . . 14327. Gene Resource Responsibilities of Federal

Agencies . . , . . . . . . . . . . . . . . . . . . . . . 15528. Estimated Economic Rates of Return From

Germplasm Accessions . . . . . . . . . . . . . . 15629. Acreage and Farm Value of Major U. S. Crops

and Extent to Which Sm all Numb ers of Varieties Dominate Crop Average . . . . . . 157

FiguresFigure No. Page

28. The Process of Plant Regeneration From

Sin gle Ce lls in Cul tu re . . . . . . . . . . . . . . . . 14729. A Model for Genetic Engineering of Forest

Trees. . . . . . . . . . . . . . . . . . . . . . . . . . . 149



138 q Impacts Applied Genetics—Micro-Organisms, Plants, and Animals

breeding objectives are specific responses to theneeds of local growers, to consumer demands,

and to the requirements of the food processing

firms and marketing systems.

Developing new varieties does the farmer lit-

tle good un less they can be in tegrated profitably

into the farming system either by increasingyields and the quality of crops or by keeping

costs down. The three major goals of crop

breeding are often interrelated. They are:

q

q

q

to maintain or increase yields by selecting

varieties for:

—pest (disease) resistance;—drought resistance;

—increased response to fertilizers; and

—tolerance to adverse soil conditions.

to increase the value of the yield by select-

ing varieties with such traits as:

—increased oil conten t;—improved storage qualities;

—improved milling and baking qualities;

an d

—increased nutritional value, such as high-

er levels of proteins.

to reduce production costs by selecting

varieties that:

—can be mechanically harvested, reducinglabor requirements;

—require fewer chemical protestants or

fertilizers; and

-can be used with minimum tillage sys-

tems, conserving fuel or labor by redu c-ing the n um ber of cultivation operations.

Th e p la n t breeder’s appr oach to

c omme rc ia l i za t ion o f ne w v ar ie t i e s

T he com m erc ia l i za t ion o f new va r i e t i e s

strongly depends on the genetic variability that

can be selected and evaluated. A typical plant

breeding system consists of six basic steps:

1. Selecting the crop to be bred.

2. Identifying the breeding goal.

3 . choosing the methodological approachneeded to reach that goal.

4. Exchanging genetic material by breeding.

5. Evaluating the resulting strain under field

conditions, and correcting any deficiencies

in meeting the b reeding goal.

6. Producing the seed for distribution to thefarmer.

The responsibilities for the different breedingphases are distributed but interactive. In theUnited States, responsibility for crop im prove-ment through plant breeding is shared by the

Federal and State governments, commercialfirms, and foundations,

2Although some specific

genes have been identified for breeding pro-grams, most imp rovements are due to gradu alselection for favorable combinations of genes insuperior lines. The ability to select promisinglines is often m ore of an ar t (involving years of experience and intuition) than a science.

The plant breeder’s approach is determinedfor the most part by the particular biologicalcharacteristics of the crop being bred—e.g., the

breeder may choose to use a system of inbreed-

ing or outbreeding, or the two in combination,as an approach to controlling and manipulating

genetic variability. The choice is influenced bywhether a particular plant in question naturally

fertilizes itself or is fertilized by a neighboring

plant. To a lesser degree, the breeding objec-

tives influence the choice of methods and the se-

quence of breeding procedures.

Repeated cycles of self-fert i l ization reducethe heterozygosity in a plant, so that after nu-

merous generations, the breeder has homozy-gous, pure lines that breed true. (See Tech. Note

4, p. 162.) Cross-fertilization, on the other hand,results in a new mixture of genes or increased

genetic variability. Using these two approaches

in combin ation produ ces a h ybrid—several lines

are inbred for homozygosity and then crossed

to produce a parental line of enhanced genetic

potential . More vigorous hybrids can be se-

lected for further testing. The effects of hybrid

vigor vary and include earlier germination, in-

creased growth rate or size, and greater crop

uniformity.

A second method for exchanging or adding

genes is achieved through altering the numberof chromosomes, or ploidy (see Tech. Note 5, p.162. ) , of the plant . S ince chromosomes are

‘National Academy of Sciences, Conservation of Germplasm Re-

sources: An Imperative, Washin gton, D . C., 1978.



Ch. 8—The Application of Genetics to Plants q 139

generally inherited in sets, plants whose ploidy

i s i nc reased usua l ly ga in fu l l s e t s o f new

chromosomes. over one-third of domesticated

spec ies a re po lyp lo ids .3General ly , crop im-

provement due to increased ploidy corresponds

to an overall enlargement in plant size; leaves

can be broader and thicker with larger flowers,fruits, or seeds. A well-known example is the

cult ivated strawberry, which has four t imes

more chromosomes than the wild type, and is

much fleshier.

Another technique, called backcrossing ca n

improve a commercially superior variety by lift-

ing one or more desirable traits from an inferior

one. Generally, this is accomplished by making a

series of crosses from the inferior to the superi-

or plant while selecting for the desired traits in

each successive generation. Self-fertilizing thelast backcrossed generation results in someprogeny that are homozygous for the genes be-ing transferred and that are identical with thesuperior variety in all other respects. Singlegene resistance to plant pests and disease-caus-ing agents has been successfully transferredthrough backcrossing.

Major cons tra in ts on crop

i m p r o v e m e n t

Two of the many constraints on crop breed-ing are related to genetics.

Many important traits are determined by several genes.

The genetic bases for imp rovements in yieldand other characteristics are not completelydefined, m ainly because most biological traits,such as plant height, are caused by the interac-tion of num erous genes. Although m any—per-haps thousands—of genes contribute to quan-titative traits, mu ch variation can be explainedby a few genes that have major impact onthe observable appearance (phenotype)

4—e.g.,

the height of some genetic dwarves in wheat

can be doubled by a single gene. Many othergenes contribute to the general health of the

3W. J. C. Lawrence, Plant Breeding [London: Edward Arnold Ltd.,

1968).

4J. N. Thompson, Jr., “Analysis of Gene Number and Develop-ment in Polygenic Systems, ” Stadler Genetics Symposium 9:63.

plant (such as resistance to pests and diseases),although some of their contributions are smalland difficult to assess. Favorable combinationsof genes result in plants well-adapted to par-ticular growing conditions and agronomic prac-tices. With th ousand s of genes in a single plant

contributing to overall fitness, the possible com-binations are almost infinite.

Most poor combinations of genes are elim-inated by selection of the best progeny; initiallyfavorable combinations are preserved and im-proved. Literally millions of plants may be ex-amined each year to find particularly favorablegenotypes for d evelopment into new breedingstocks. Increasingly sophisticated field testingprocedures, as well as advanced statistical anal-yses, are now used to evaluate the success of breeding efforts. Overall yield is still the mostimportant criterion for success, although con-siderable care is taken to test stress tolerance,pest and disease resistances, mechanical har-vestability, and consumer acceptability. Breed-ing programs with specialized goals often userapid and accurate chemical procedures toscreen lines and p rogeny for imp rovements.

Because the vigor of the plant d epend s on theinteraction of many genes, it has been difficultto identify individual genes of physiologicalsignificance in w hole plants. As a result, manyimportant genes have not been mapped inmajor crop species. There is little doubt that

breeders w ould select traits like photosyn theticefficiency (the ability to convert light to suchorganic compounds as carbohydrates) or miner-al uptake if the genes could be identified andmanipulated in the same ways that resistance isselected for pathogens.

It is uncertain how much genetic variation for improvement exists.

Although the world’s germplasm resourceshave not been completely exploited, it hasbecome m ore difficult for breeders to improvemany of the highly developed varieties now inuse-e. g., height reduction in wheat has madeenormou s contributions to its produ ctivity, butfurther improvement on this basis seems to belimited.

5A par allel cond ition in the pota to crop

‘N. ~. Jensen, “Limits to Growth in World Food Product ion,” Sci-

ence 201:317, 1978.



Ch. 8—The Application of Genetics to Plants . 141

New genetic technologies for

p lan t b re e d ing

The recent breakthroughs in genetic engi-neering p ermit the plant breeder to byp ass thevarious natural breeding barriers that havelimited control of the transfer of genetic in-formation. While the new technologies do notnecessarily offer the plant breeder the radicalchanges that recombinant DNA (rDNA) technol-ogy provides the microbiologist, they will, intheory, speedup and perfect the process of ge-netic refinement.

The new technologies fall into two catego-ries: those involving genetic transformationsthrou gh cell fusion, and those involving the in-sertion or modification of genetic informationthrou gh the cloning (exactly copying) of DNAand DNA vectors (transfer DNA). Most genetic

transformations require that enzymes digestthe plant’s impermeable cell wall, a process thatleaves behind a cell without a w all, or a p roto-plasm. Protop lasts can fuse w ith each other, aswell as with other components of cells. Intheory, their ability to do this permits a widerexchange of genetic information.

The approach exploiting the new technol-ogies is usually a three-phase program ..

Phase 1. Isolated cells from a plant are estab-lished in tissue culture and kept alive.

Phase 11. Genetic changes are engineered inthose cells to alter the genetic makeupof the plant; and desired traits areselected at th is stage, if possible.

Phase 111. The regeneration of the altered singlecells is initiated so that they grow intoentire plants.

This approach contains similarities to the genet-ic manipu lation of micro-organisms. However,there is one major conceptual difference. Inmicro-organisms, the changes m ade on the cel-lular level are the goals of the manipulation.With crops, changes mad e on the cellular level

are meaningless unless they can be reprodu cedin the entire plant. Therefore, unless single cellsin culture can be grown into mature plants thathave th e new , desired characteristics—a proce-dure which, at this time, has had limited suc-cess—the benefits of genetic engineering will

not be widespread. If the barriers can be over-come, the new technologies will offer a newway to control and d irect the genetic character-istics of plants.

PHASE 1: TISSUE CULTURE TO CLONE PLANTS

Tissue cultur e involves grow ing cells from a

plant in a culture or medium th at will sup portthem an d keep th em viable. It can be started atthree different levels of biological organization:with plant organs (functional units such asleaves or roots);* with tissues (functioning ag-gregates of one typ e of cell, such as ep idermalcells (ou term ost layer) in a leaf; and with singlecells. Tissue cultures by themselves offer spe-cific benefits to plant breeders; just as fermenta-tion is crucial to microbial genetic technologies,tissue culture is basic to the application of theother new genetic technologies for plants.

The idea of growing cells from higher p lantsor animals and then regenerating entire plantsfrom these laboratory-grown cells is not new.However, a better scientific understanding nowexists of wh at is needed to keep the plant p artsalive.

In tissue culture, isolated single plant cells aretypically induced to undergo repeated cell divi-sions in a broth or gel, the resulting amorphouscell clump is known as a callus. If culture condi-tions are readjusted when the callus appears, itscells can undergo further proliferation. As the

resulting cells differentiate (become special-ized), they can grow into the well-organized

tissues and organs of a complete normal plant.The callus can be fur ther su bcultured, allowingmass pr opagation of a desired plant.

At this time, it is not un common to p rodu ce asmany as a thousand plants from each gram of starting cells; 1 g of starting carrot callus rou-tinely produces 500 plants. The ultimate goal of tissue culturing is to have these plantlets placedin regular soil so that they can grow and devel-op into fully functional mature plants. The com-plete cycle (from plant to cell to plant) permitsproduction of plants on a far more massivescale, and in a far shorter period, than is possi-ble by conventional means. (See table 25 for a

*Also referred to as organ culture.




First stage in plant tissue culturing: inoculationof plant tissue

cies basis. However, several commercial uses of tissue cultu re alread y exist. (See table 26.)

Storage of Germplasm.—Tissue culturecan be used in the long-term storage of special-ized germ plasm, w hich involves freezing cells

and types of shoots. The culture p rovides stablegenetic material, reduces storage space, anddecreases maintenance costs.

Shows the gradual development of the plant t issueon an agar medium

list of some pculture. )

ants propagated through tissue

Each of the four stages of the completecycle—establishment in culture, organogenesis,plantlet amplification, and reestablishment insoil—requires precise biological environmentsthat have to be determined on a species-by-spe-

Carrot tissues have been frozen in liquid ni-trogen, thawed 2 years later, and regeneratedinto normal plants. This technique has alsoproved successful with morning glories, syca-mores, potatoes, and carnations. Generally, thetechn ique is most useful for plant m aterial thatis vegetatively propagated, although if it can begenerally ap plied it could become importan t forother agriculturally important crops.

P r o d u c t i o n o f p h a r m a c e u t i c a l s a n dOther Chemicals From Plant Cells.—Be-cause plant cells in culture are similar to micro-organisms in fermentation systems, they can beengineered to work as “factories” to produce




Table 25.—Some Plants Propagated ThroughTissue Culture for Production or Breeding

Agriculture and Poinsettiahorticulture Weeping fig

Vegetable crops Rubber plantAsparagus FlowersBeet African violet

Brussels sprouts AnthruiumCauliflower ChrysanthemumEggplant Gerbera daisyOnion GloxiniaSpinach PetuniaSweet potato RoseTomato Orchid

Fruit and nut trees FernsAlmond Australian tree fernApple Boston fernBanana Maidenhair fernCoffee Rabbitsfoot fernDate Staghorn fernGrapefruit Sword fernLemon BulbsOlive LilyOrange Daylily

Peach Easter lilyFruit and berries Hyacinth

BlackberryGrape

PharmaceuticalAtropa

PineappleStrawberry

GinsengPyrethium

FoliageSilver vase Silviculture (forestry)

Begonia Douglas fir

Cryptanthus Pine

Dieffenbachia Quaking aspen

Dracaena Redwood

Fiddleleaf Rubber tree


Table 26.—Representative List of Tissue Culture Programs of Commercial Significance in the United States

Industry Application Economic benefits

Asparagus industry. . . . . . . . . .Chemical and pharmaceutical.

Citrus industry. . . . . . . . . . . . . .Coffee industry . . . . . . . . . . . . .Land reclamation. . . . . . . . . . . .Ornamental horticulture. . . . . .

Pineapple industry . . . . . . . . . .

Strawberry industry. . . . . . . . . .

Rapid multiplication of seed stock Improved productivity, earliness, and spear qualityBiosynthesis of chemicals Reduced production costsPropagation of medicinal plants High volumes of plants for plantingVirus elimination Improved quality, high productivityDisease resistance breeding Disease resistanceMass propagation Availability of select clones of wild species for revegetationMass propagation Reduced costs of certain species

Virus elimination of certain speciesIntroduction of new selectionsIncreased volumes of difficult selections

Mass propagation Improved quality in higher volumes

Mass propagation Rapid introduction of new strains





plant products or byproducts. In recent years,economic benefits have been achieved from theproduction of plant constituents through cellculture. Among those currently produced com-mercially are camptothecin (an alkaloid withantitumor and antileukemic activity), pr oteinaseinhibitors (such as hep arin), and antiviral sub-stances. Flavorings, oils, other medicinal, andinsecticides will also probably be extracted fromthe cells.

The vinca alkaloids—vincristine and vin-blastine, for instance—are major chemothera-peutic agents in the treatment of leukemias andlymphomas. They are derived from the leavesof the Madagascar periwinkle (Catharanthus roseus). Over 2,000 kilograms (kg) of leaves arerequired for the production of every gram of vinca alkaloid at a cost of about $250/ g. Plantcells have recently been isolated from the peri-

winkle, imm obilized, and placed in culture. Thisculture of cells not only continues to synthesizealkaloids at high rates, but even secretes the ma-terial directly into the cultu re med ium insteadof accumu lating it within the cell, thu s remov-ing the need for extensive extraction pro-cedures.

Similarly, cells from the Cowage velvetbeanare currently being cultured in Japan as asource of L-Dopa, an important drug in the

treatment of Parkinson’s disease. Cells from the

opium popp y synthesize both the p lant’s normal

alkaloids in culture and, apparently, some alka-loids that have n ot as yet been foun d in extracts

from the whole plant.

Another pharmaceutical, diosgenein, is the

major raw material for the production of corti-

costeroids and sex steroids like the estrogens

and progestins used in birth control pill. Thel ar ge tu b e ro u s r o ot s o f it s p l an t s o u r ce ,

Dioscorea, are still collected for this purpose inthe jungles of Central Am erica, but its cells havebeen cultured in th e laboratory.

Other plant products, from flavorings and

oils to insecticides, industrial organic chemicals,and sweeteners, are also beginning to be de-rived from plants in cell-cultures. Glycyrrhiza,the nonnutritive sweetener of licorice, has beenprodu ced in cultures of Glycyrrhiza glabra, and

anthraquinones, which are used as dye bases,accumulate in copious amounts over severalweeks in cultures of the mulberry, Morinda citri-

folia.

PHASE II: ENG INEERING CHANG ES TO ALTERGENETIC MAKEUP; SELECTING DESIRED TRAITS

The second phase of the cycle involves thegenetic manipu lation of cells in tissue culture,followed by the selection of desired traits,Tissue culturing, in combination w ith the newgenetic tools, could allow the insertion of newgenetic information directly into plant cells.Several approaches to exchanging genetic infor-mation through new engineering technologiesexist:

q culturing plant sex cells and embryos;q protoplasm fusion; andq transfer by DNA clones and foreign

tors.

These are then followed b y:

. screening for desired traits.

vec-

Cul tur ing P lan t Ce l l s and Embryos .–Culturing the plant’s sex cells—the egg from theovary and the pollen from the anther (pollen-secreting organ)—can increase the efficiency of creating pure plant lines for breeding. Since sexcells contain only a single set of unpairedchromosomes per cell, plantlets derived fromthem also contain only a single set. Thus, anygenetic change will become ap paren t in the re-

generated plant, because a second paired genecannot mask its effect. Large numbers of hap-loid plants (cells contain half the norm al num -ber of chromosomes) have been produced formore than 20 species. Simple treatment withthe chemical, colchicine, can usually inducethem to duplicate their genomes (haploid set of chromosomes) —resulting in fully normal, dip-loid plants. The only major crop that h as beenbred by th is technique is the asparagus.

8

If the remaining technical barriers can beovercome, the technique can be used to en-

hance the selection of elite trees and to createhybrids of important crops. Although still

‘J . G. Tor r eey , “Cytodifferentiation in Cultured Cells and Tissues, ”l-/ortScience 12(2):138, 1977.




primarily experimental, successful plant sex-cellcultures have been achieved for a variety of important cultivars, including rice, tobacco,wheat, barley, oats, sorghum, and tomato. How-ever, because the techn ique can lead to bizarreunstable chromosomal arrangements, it has had

few applications.

Embryo cultures have been used to germi-nate, in vitro, those embryos that might nototherwise survive because of basic incompatibil-ities, especially when plants from differentgenera are crossed. Embryos may function asstarting material in tissue culture systems re-quiring juven ile material. They are being u sedto speed up germination in such species as oilpalms, which take up to 2 years to germinateunder natural conditions.

P r o t o p l a s m F u s i o n . —In protoplasm fusion,

either two entire protoplasts are brought to-gether, or a single protoplasm is joined to cell

components —or organelles —from a second pro-toplasm. When the comp onents are mixed u nderthe right cond itions, they fuse to form a singlehybrid cell. The hybrids can be indu ced to pro-liferate and to regenerate cell walls. The func-tional plant cell that results may often becultured further an d regenerated into an entireplant—one that contains a combination of genet-ic material from both starting plant cell progeni-tors. When protoplasts are indu ced to fuse, they

can, in theory, exchange genetic informationwithou t the restriction of natu ral breeding bar-riers. At present, protoplasm fusion still hasman y limitations, mainly d ue to the instability of chromosome pairing.

Organelles are small, specialized componentswithin the cell, such as chloroplasts and mito-chondria. Some organelles, called plastids, carrytheir own autonomously replicating genes, as aresult, they may hold p romise for gene transfer

and for carrying new genetic information intoprotoplasts in cultures, or possibly for influenc-ing the functions of genes in the cell nucleus.

(See Tech. N ote 8, p . 163.)

The feasibility of protoplasm fusion has beenborne out in recent work with tobacco–a plantthat seems particularly amenable to manipula-tion in culture. An albino mutant of Nicotiana

tabacum was fused with a variety of a sexuallyincompatible Nicotiana species. The resultanthybrids were easily recognized by their inter-med iate light green color. They have now beenregenerated into adult plants, and are currentlybeing used as a prom ising sour ce of hornworm

resistance in tobacco plants.

Transfer by DNA Clones and ForeignVectors.—Recombinant DNA technologymakes possible the selection and prod uction of more copies (amplification) of specific DNAsegments. Several basic approaches exist. In the“shotgun” approach, the whole plant genome iscut by one or more of the commercially avail-able restriction enzym es. The DN A to be tran s-ferred is then attached to a plasmid or phage,which carries genetic information into the plantcell.—E.g., a gene coding for a protein (zein) that

is a major component of corn seeds has beenspliced into p lasmids and cloned in micro-orga-nisms. It is hoped that the zein-gene sequencecan be modified through this approach to in-crease the nutritional quality of corn proteinbefore it is reintrodu ced into the corn plant.

Foreign vectors are nonplant materials (vi-ruses and bacterial plasmids) that can be used totransfer DNA into higher plant cells. Trans-formation through foreign vectors might im-prove plant varieties or, by amplifying the de-

sired DNA sequence, make it easier to recover acell prod uct from culture. In ad dition, methodshave been discovered that eliminate the foreignDNA from th e transformed mixture, leavingonly the desired gene in the transformed plant.The most promising vector so far seems to bethe tumor-inducing (Ti) plasmid carried by

Agrobacterium tumefaciens. This bacteriumcauses tumorous growths around the rootcrowns of plan ts. It infects one m ajor grou p of plants–the d icots (such as peas and beans), so-called because th eir germinating seeds initiallysprou t dou ble leaves. Its virulence is du e to the

Ti plasmid, which, when it is transferred toplant cells, induces tumors. Once inside the cell,a smaller segment of the Ti plasmid, called T-DNA, is actually incorporated into the recipientplant cell’s chromosomes. It is carried in thisform, replicating right along with the rest of the




chromosomal DNA as plant cell proliferationproceeds. Researchers have been wonderingwhether new genetic material for plant im -provement can be inserted into the T-DNA

region and carried into plant cell chromosomes

in functional form.

Addin g foreign genetic material to the T-DNAregion has proved successful in several ex-

periments. Furthermore, it has been found thatone type o f p l an t t um or ce l l t ha t con ta ins

mutagenized T-DNA can be regenerated into acomplete plant . This new discovery supportsthe use of the Agrobacterium system as a modelfor the introduction of foreign genes into the

single cells of higher p lants.

Many unanswered questions remain before Agrobacterium becomes a useful vector forplant breeding. Considerable controversy existsabout exactly where the Ti plasmid integratesinto the host plant chromosomes; some inser-tions might disrupt plant genes required forgrowth. In addition, these transformations maynot be genetically stable in recipient plants;there is evidence that the p rogeny of Ti-plasmid-containing plants d o not retain copies of the Tisequence. Finally, Agrobacterium does not read-ily infect m onocots (a second group of plants),which limits its use for major grain crops.

Another prom ising vector is the cauliflowermosaic virus (CaMV). Since none of the knownplant DNA viruses has ever been found in p lant

nu clear DNA, CaMV may be u sed as a vector forin t roducing gene ti c in form at ion in to p lan t

cytoplasm. Although studies of the structuralorganization, transcription, and translation of the CaMV are being undertaken, information

available today suggests that the system needsfurther evaluation before it can be considered

an alternative to the Agrobacterium system.

Although work remains to be done on Ti-plasmid and CaMV genetic mechanisms, thesesystems have enormou s potential. Most immedi-ately, they offer ways of examining basic mech-anisms of differentiation and genetic regulationand of delineating the organization of thegenome within the h igher p lant cell. If this canbe accomplished, the systems may provide away of incorpor ating complex genetic traits intowhole plants in stable and lasting form.

Screening for Desired Traits.—The bene-fits of any genetic alteration will be realizedonly if they are combined with an adequate sys-tem of selection to recover the desired traits. Insome cases, selection pressures can be useful inrecovery.

gThe toxin from plant p athogens, for

example, can help to identify disease resistancein plants by k illing th ose that are n ot resistant.So far, this method has been limited to id entify-ing toxins excreted by bacteria or fungi andtheir an alog; after sugarcane calluses w ere ex-posed to toxins of leaf blight, the resistant linesthat survived were then used to develop newcommercial varieties. In theory, h owever, it ispossible to select for many important traits.Tissue culture breeding for resistance to salts,herbicides, high or low tempera tures, drou ght,and new varieties that are m ore responsive tofertilizers is currently under study.

Five basic problems mu st be overcome beforeany selected trait can be considered beneficial(see figure 28):

the trait itself mu st be identified;a selection scheme m ust be found to iden-tify cells w ith altered pr operties;the properties must prove to be du e to ge-netic changes;cells with altered properties must confersimilar pr operties on the wh ole plant; andthe alteration must not adversely affectsuch commercially important characteris-tics as yield.

While initial screens involving are easier tocarry out than screening tests involving entireplants, tolerance at the cellular level must beconfirm ed by inoculations of the matu re plantswith the actual pathogen u nd er field conditions.

PHASE III: REGENERATING WHOLE PLANTSFROM CELLS IN TISSUE CULTURE

New methods are being developed to:

q increase the speed with which crops are

multiplied through mass propagation, and. create and maintain disease-free plants.

Mass Propagation.—The greatest singleuse of tissue culture systems to date has beenfor mass propagation, to establish selected

‘J. F. Shepa rd , D. Bidney, and E. Shahin , “potato Pr o to p ]a s t s in

Crop Imp rovement,” Science 208:17, 1980.




Figure 28.—The Process of Plant Regeneration From Single Cells in Culture

Virus-free

Field performance tests

Cell multiplication

Tissue

Cell wallremoval

+ Root-promotinghormones

The process of plant propagation from single ceils in culture can produce plants with selected characteristics. These selec-tions must be tested in the field to evaluate their performance.


Photo credit: Plant Resources Institute

Multiplying shoots of jojoba plant in tissue culture on apetri dish. These plants may potentiality be selected for

higher oil content

culture because of the increased speed with

sources of improved seed or cutting material.(See table 26.) In some cases, producing plantsby other means is simply not economically com-

petitive. A classic example is the Boston fern,

which, while it is easy to propagate from runner

tips, is commercially propagated through tissue

which i t multiplies and the reduced costs of

stock plant maintenance. A tissue culture stock

of only 2 square feet (ft2) can produce 20,000

plants per month.10

Currently, mass production of such cultivars

as strawberries (see Tech. Note 9, p. 163.),asparagus, oil palms, and pineapples is beingcarried ou t through plant tissue cultures.

11Very

recently, alfalfa was propagated in the same

way, giving rise to over 200,000 plants, several

thousand of which are currently being tested in

field trials. Also, 1,300 oil palms, selected for

high yield and disease resistance, are being

tested in Malaysia. 12 Other crops not produced

by this method but for which cell culture is an

important source of breeding variation include

‘OD. P. Holdgate, ‘(Propagation of Ornamental by Tissue Cul-ture, ” in Plant Cell, Tissue, and Organ C’ulture, .1. Feinert and Y. P. S.

Bajaj (eds.) (New’ York: Sprin@r-\rerla~, 1977).

“T . Murashige, “Current Status of Plant Cell and organ Cul-

tures, ” FfortScience )2(2):127, 1977.

‘z

’’ The Second Green Revolution, ” special report, Business Week,

/W~. 25, 1980.




beets, brussels sprout, cauliflower, tomatoes,

citrus fruits, and bananas. Various horticultural

plants—such as chrysanthemums, carnations,

African violets, foliage plants, and ferns—are

also being produ ced b y in vitro techn iques.

Accelerating propagation and selection in

culture is especially compelling for economical-

ly important forest species for which traditional

breeding approaches take a century or more.

Trees that reach maturi ty within 5 years re-

quire approximately 50 years to achieve a usefulhomozygous strain for further breeding. Spe-

cies such as the sequoia, which do not floweruntil they are 15 to 20 years old, require be-tween 1 and 2 centuries before traits are sta-bilized and preliminary field trials are eval-

uated. Thus, tissue culture production of treeshas become an area of considerable interest.Already, 2,500 tissue-cultured redwoods havebeen grown under field conditions for compari-son with regular, sexually produced seedlings.(See ap p. II-B.) Loblolly p ine an d Douglas fir are

also being cultured; the number of trees thatcan be grow n from cells in 100 liters (]) of med iain 3 months are enough to reforest roughly120,000 acres of land at a 12 x 12 ft spacing.

13To

date, 3,000 tissue-cultured Douglas firs have ac-tually been planted in natural soil conditions.(See figu re 29.)

1 3D . J . Durzan, “Progress and Promise in Forest Genetics,” inProceedings, 50th Anniversary Symposium Paper, Science and

Technology. . . The Cutting Edge (Appleton, Wis: Institute of PaperChemistry, 1980).

A plantlet of Ioblolly pine grown in Weyerhaeuser Co.’stissue culture laboratory. The next step in this procedure

is to transfer the plant let from its sterile and humidenvironment to the soil




Figure 29.—A Model for Genetic Engineeringof Forest Trees

virus-free stock often ap pear a s larger flowers,more vigorous growth, and improved foliagequality.

a.

f.

a.

b.

c.

d.

e.

Selection of genetic material from germplasm bank

Insertion of selected genes into protoplasts

Regeneration of cells from protoplasts and

multiplication of cell clones

Mass production of embryos from cells

Encapsulation to form ‘seeds’

f. Field germination of ‘seeds’

g. Forests of new trees


C r e a t i o n a n d M a i n t e n a n c e o f D i s e a s e -

Free Plants.—Cultivars maintained throughstandard asexual p ropagation over long p eriods

often pick up viruses or other harmful path-ogens, which while they might not necessarily

kill the plants, may cause less healthy growth. Aplant’s true economic potential may be reached

only if these pathogens are removed—a task

which culturing of a plant’s meristem (growingpoint ) and subsequent heat therapy can per-

form. Not al l plants produced through these

methods are virus-free, so screening cells for

viruses must be done to ensure a pathogen-free

plant. In horticultural sp ecies, the adv antages of

Today, virus-free fruit plants are m aintainedand distributed from both private and publicrepositories. Work of commercial importance

has been done with such plants as strawberries,sweet potatoes, citrus, freesias, irises, rhubarbs,gooseberries, lilies, hops, gladiolus, geraniums,and chrysanthemums. 14 Over 134 virus-freepotato cultures have also been developed by tis-sue culture.

15

C ons t ra in t s on th e ne w gene t i c

t e c hno log ie s

Although genetic information has been trans-ferred by vectors and protoplasm fusion, no DNAtransformations of commercial value have yetbeen performed. The constraints on the suc-cessful application of molecular genetic technol-ogies are both technical and institutional.

TECHNICAL CONSTRAINTS

Molecular engineering has been imped ed by alack of understand ing about wh ich genes wou ldbe useful for plant breed ing pu rposes, as well asby insufficient knowledge about cytogenetics.In addition, the available tools—vectors andmu tants—and methods for transforming plantcells using pu rified DN A are still limited.

Cells carrying traits important to crop pro-ductivity must be identified after they havebeen genetically altered. Even if selection for anidentified trait is successful, it must be dem-onstrated that cells with altered properties con-fer similar properties on tissues, organs, and,ultimately on the whole plant, and that thegenetic change does not adversely affect yieldor other desired characteristics. Finally, onlylimited success has been achieved in regenerat-ing whole plants from individual cells. While thelist of plant species that can be regeneratedfrom tissue culture has increased over the last 5

years, it includes mostly vegetables, fruit and

lqh~. NI isa~~,a, K. Sak~IO, hl. ‘ 1 ’ i i n a k a , N1 . Havashi, an d H . SaIII~-

jima, “Production of Physiologically Act i \ w Substances by Plant(kll Suspension (cultures, ” H. E. SIIYXX (cd.), Tissue Culture an d

Plant Science (New York: Academic Press, 1974).jshlul,ashige, op . Cit .

76-565 0 - 81 - 11



150 “ Impacts of Applied Genetics —Micro-Organisms, Plants, and Animals

nut trees, flowers, and foliage crops. Some of the most important crops—like w heat, oats, andbarley–have yet to be regenerated . In ad dition,cells that form calluses in culture cannot alwaysbe coaxed into forming embryos, which mustprecede the formation of leaves, shoots, and

roots. Techn ical breakthroughs have come on aspecies-by-species basis; key technical discov-eries are not often ap plicable to all plants. Andeven when the new technologies succeed intransferring genetic information, the changescan be u nstable.

The hope that protoplasm fusion would openextensive avenues for gene transfer betweendistantly related plant species has diminishedwith the observation of this instability. How-ever, if whole chromosomes or chromosomefragments could be transferred in plants wheresexual hybridization is presently impossible, thepossibilities would be enorm ous.

INSTITUTIONAL CONSTRAINTS

Institutional constraints on m olecular gen et-ics include those in funding, in regulation, inmanpow er, and in indu stry.

Federal fund ing for plant m olecular geneticsin agriculture has come from the NationalScience Found ation (N SF) and from USDA. Re-search support in USDA is channeled primarilythrough the flexible Competitive Grants Pro-gram (fiscal year 1980 budget of $15 million) for

the sup port of new research directions in plantbiology. The pan el on genetic mechanisms (an-nual budget less than $4 million) is of particularsignificance for developing new genetic technol-ogies. The panel’s charter specifically seeks pro-posals on novel genetic technologies. The re-maining three panels concerned with plants—nitrogen, photosyn thesis, and stress—also sup-port projects to define the molecular basis of fundamental plant properties. The success of the USDA Competitive Grants Program is hardto assess after just 2 years of operation; how-ever, its bud get over the past 2 years has severe-

ly limited expansion of the program into newareas of research.

Some private institutions16

argue that the

W. walbo t , past, present and Future Trends in Cm p f3medin&

Vol. H, Working Papers, Impact o~appiied Genetics,NTI S, 1981.

Competitive Grants Program is shifting supportfrom ongoing USDA prog rams to new geneticsresearch programs that are not aimed at theimportant problems facing agriculture today.There is no opposition to supporting the molec-ular approaches as long as they do not come at

the expense of traditional breeding programs,and as long as both molecular biologists andclassical geneticists working with major cropplants are assured of enough sup port to fosterresearch groups of sufficient size.

At present, fund s from n ine programs at theNSF—primarily in the Directorate for Biologic],Behavioral, and Social Sciences—support plantresearch. The total support for the plant sci-ences may be as high as $25 million, of whichonly about $1 million is designated specificallyfor molecular genetics.

The regulation of the release of genetically altered plants into the environment has not hadmuch effect to date. As of November 1980, onlyone application [which requested exceptionfrom the NIH Guidelines (see ch. 11) to releaserDNA-treated corn into the environment] hasbeen filed with the Office of Recombinant DNAActivities (ORDA). Whether regulation will pro-duce major obstacles is difficult to predict atpresent. It is also unclear whether restrictionswill be placed on other genetic activities, suchas protoplasm fusion. Currently, at least oneother nation (New Zealand) includes such re-

strictions in its guidelines. It is not clear howmuch the uncertainty of possible ecological dis-ruption and the attendent liability concernsfrom intentional release of genetically engi-neered plants has prevented the industrial sec-tor from moving tow ard commercial applicationof the new technology.

Only a few universities have expertise in bothplant an d molecular biology. In ad dition, only afew scientists work with modern moleculartechniques related to whole plant problems. Asa result, a business firm could easily develop a

capability exceeding that at any ind ividual U.Suniversity. However, building industrial laboratories and hiring from the universities couldeasily deplete the expertise at the universitylevel. With the recent investment activity inbioengineering firms, this trend has already




begun; in the long-run it could have serious con-

sequences for the quality of university research.

Despite these constraints, progress in over-

coming the diff icult ies is continuing. At theprestigious 1980 Gordon Conference, where sci-

entists meet to exchange ideas and recent find-

ings, plant molecular biology was added to the

list of meetings for the first time. In addition,

four other recent meetings have concentrated

on plant molecular biology.17

Up to 50 percent

of the participants at these meetings came fromnonplant-oriented disciplines searching for fu-

ture research topics. This influx of investigators

from other fields can be expected to enrich the

variety of approaches used to solve the prob-

lems of the plant breeder.

I TGenome Organization and Expression in Plants, NATO s.vm-

posium held in Edinburgh, Scotland, July 1979; Genetic Engineering

of Symbiotic Nitrogen Fi&ation and Conservation of Fi~ed Nitrogen t June 29-Juiy 2, 1980, Tahoe City, Ca]if; ‘“Molecular Biologists Look al Green Plants,” Si,xfh Annual Symposium, Sepl: 29-oct. 2, lg80,}k?idelberg, West [;ermany; and Fourth /nferna[ional Symposium

on Nitro,qm F~xation, Dec. 1-.5, 1980, Canberra, Aust ral iii.

Finally, as a general rule, tradeoffs arise inthe use of the new technologies that may inter-fere with their application. It is impossible to getsomething for noth ing from nature—e.g., in ni-trogen fixation the symbiotic relationship bet-ween plant and micro-organism requires ener-

gy from the plant; screening for plants th at canprodu ce and transfer the end p rodu cts of pho-tosynthesis to the nodu les in the root mor e effi-ciently may r edu ce inorgan ic nitrogen requ ire-ments but may also reduce the overall yield.This was the case for the high lysine varieties of corn. (See Tech. Note 10, p. 163.) Farm ers in theUnited States tended to avoid them because im-proving the protein quality reduced the yield,an unacceptable tradeoff at the market price.Thus, unless the genetic innovation fits the re-quirements of the total agricultural industry,potentials for crop improvement may not be

realized.

I m p a c t s on g e n e r a t in g n e w v a r ie t ie s

Progress in the manipulation of gene expres-sion in eukaryotic (nucleus-containing) cells,which include the cells of higher plants, hasbeen enormous. Most of the new methodologieshave been derived from fruit flies and mam-

malian tissue cultu re lines; but m any shou ld bedirectly app licable to studies with p lant genes.There has been great progress in isolating spe-cific RNA from plants, in cloning plant DNA, andin un derstanding m ore about the organizationof plant genomes. Techniqu es are available formanipulating organs, tissues, cells, or pro-toplasts in culture; for selecting markers; forregenerating p lants; and for testing the geneticbasis of novel traits. So far however, thesetechniques are routine only in a few species.Perfecting procedures for regenerating singlecells into w hole plants is a p rerequisite for thesuccess of many of the novel genetic technol-ogies. In addition, work is progressing onviruses, the Ti plasmid of Agrobacterium, andengineered cloning vehicles for introducingDNA into plants in a directed fashion. There

have been few demonstrations in which the in-heritance of a new trait was maintained overseveral sexual generations in the w hole plant.

Because new varieties have to be tested

under different environmental conditions oncethe problems of plant regeneration are over-come, it is difficult to assess the specific impactsof the new technologies.—E. g., it is impossible todetermine at this time whether technical andbiological barriers will ever be overcome forregenerating wheat from protoplasts. Never-theless, the imp act of genetics on th e structureof American agr icultu re can be d iscussed withsome degree of confidence.

Genetic engineering can affect not only w hatcrops can be grown, but wh ere and how thosecrops are cultivated. Although it is a variable inproduction, it usually acts in conjunction withother biological and mechanical innovations,whose deployment is governed by social, eco-nom ic, and political factors.




testing is needed to determine wh ether the improvementcan be maintained in field trials, where the improvedstrains must compete against wild-type Rhizobia alreadypresent in the soil.

Another w ay to imp rove nitrogen fixation is to selectplants th at have more efficient symb iotic relationship swith nitrogen-fixing organism s. Since the biological proc-

ess requires a large amount of en ergy from the plan t, itmay be p ossible to select for plants that are more efficientin produ cing, and then to transfer the end p roducts of photosynthesis to the nodules in the roots. Also existingnitrogen-fixing b acterial strains th at can interact with cropplants which do not ordinarily fix nitrogen could besearched for or developed.

Reducing the amount of chemically fixed nitrogenfertilizer–and the cost of the natural gas previously usedin the chemical process—would be the largest benefit of successfully fixing nitrogen in crops. Environmental bene-fits, from the smaller amount of fertilizer runoff intowater systems, would accrue as w ell. But is it d ifficult topredict when these will become reality. Experts in the fielddisagree: some feel the breakthrou gh is immin ent; othersfeel that it might take several decades to achieve.

The refinements in breeding methods pro-vided by the new technologies may a llow m ajorcrops to be bred more and more for specializeduses—as feed for specific animals, perhaps, orto conform to special processing requ irements.In add ition, since the popu lations in less devel-oped countries suffer more often from majornutritional deficiencies than those in industrial-ized countries, a specific export market of cere-al grains for human consumption, like wheatwith higher p rotein levels, may be d eveloped.

But genetic methods are only the tools andcatalysts for the changes in how society pro-du ces its food; financial pressur es and Federalregulation will continue to direct their course.E.g., the automation of tissue culture systemswill decrease the labor needed to direct plantpropagation and drastically reduce the cost perplantlet to a level comp etitive w ith seed p ricesfor many crops. While such breakthroughs mayincrease the commercial applications of manytechnologies, the effects of a displaced laborforce and cheaper and more efficient plants arehard to pr edict.

Although it is difficult to make economic projections, there are several areas wh ere genetictechnologies w ill clearly have an impact if thepredicted breakthroughs occur:

q

q

q

q

For

Batch culture of plant cells in automatedsystems w ill be enhan ced by the ability toengineer and select strains that producelarger quantities of plant substan ces, suchas pharmaceutical drugs.The technologies w ill allow developm ent of elite tree lines that will greatly increaseyield, both through breeding programssimilar to those used for agricultural cropsand by overcoming breeding barriers andlengthy br eeding cycles. Refined method sof selection and hybridization will increasethe potential of short-rotation forestry,which can provide cellulosic substrates forsuch prod ucts as ethanol or methanol.The biological efficiency of many economi-cally important crops will increase. Ad-vances will depend on the ability of thetechniques to select for whole plant charac-teristics, such as photosynthetic soil andnutrient efficiency.

19

Besides narrowing breeding goals, thetechniques will increase the potential forfaster improvement of underexploitedplants w ith promising economic value.

such advances to occur, genetic factorsmust be selected from superior germplasm, the

genetic contributions must be integrated intoimproved cultural practices, and the improvedvarieties must be efficiently propagated fordistribution.

For the soybean and tomato crops, the research area for im-

proved biological efficiency received the highest allotment of

funds in fiscal year 1978. Total fun din g was $12.9 million for so y-beans and $2.1 million for tomatoes. The second largest categoryto be fun ded w as control of diseases and n ematodes of soybeans at

$5.1 mill ion and for tomato at $1.6 mill ion.




G e n e t i c v a r i a b i l i t y , c r o p v u l n e r a b i l i t y , a n d

s t o r a g e o f g e r m p l a s m

Successful plant breeding is based on theamount of genetic diversity available for the in-

sertion of new genes into plants. Hence, it isessential to have an adequate scientific under-standing of how m uch genetic erosion has takenplace and how much germplasm is needed. Nei-ther of th ese questions can be satisfactorily an-swered today.

T he amoun t o f ge ne t i c e ros ion tha t

h a s t a k e n p l a ce

Most genetic diversity is being lost because of the d isplacement of vegetation in areas outsidethe United States. The demand for increasedagricultural production is a principal pressurecausing d eforestation of trop ical latitud es (seeTech. Note 11, p. 163), zones that contain exten-sive genetic diversity for both plants andanimals.

It has been estimated that several hundredplant species become extinct every year andthat thousands of indigenous crop varieties(wild types) have alread y been lost. How ever, itis difficult to measure this loss, not only becauseresources are on foreign soil but because ero-sion mu st be examined on a species-by-species

basis. In theory, an adequate evaluation wou ldrequire knowledge of both the quantity of di-versity within a species and th e breadth of thatdiversity; this process has in practice, just be-gun. What is known is that the lost material can-not be replaced.

T h e a m o u n t o f g e r m p l a s m n e e d e d

Germplasm is needed as a resource for im-proving characteristics of plants and as a m eansfor guaranteeing supplies of known plantderivatives and potential new ones. Even if

plant breeders adequately understand theamou nt of germplasm p resently needed , it is dif-

ficult to predict future needs. Because pests andpathogens are constantly mutating, there is

always the possibility tha t some resistance willbe broken dow n. Even thou gh genetic diversitycan reduce the severity of economic loss, an epi-demic might require the introduction of a newresistant variety. In addition, other pressureswill determine which crops will be grown forfood, fiber, fuel, and ph armaceuticals, and howthey w ill be cultivated; genetic diversity will befundamental to these innovations.

Even if genetic needs can be ad equately iden-tified, there is disagreement about how much

germplasm to collect. In the past, its collectionhas been guided by differences in morphology(form and structure), which have n ot often beendirectly correlated to breeding objectives. Fur-thermore, the extent to which the new genetictechnologies will affect genetic variability, vul-nerability, or the storage of germplasm, has notbeen d etermined. (See app . II-A.)

In add ition to its uses in plant improvem ent,germplasm can provide both old and new p rod-ucts. Recent interest in growing guayule as asource of hydrocarbons (for rubber, energy

materials, etc.) has focused atten tion on plantsthat m ay possibly be und erutilized. It has beenfound that past collections of guayule germ-plasm have not been adequately maintained,making current genetic improvements more dif-ficult. In addition, half of the w orld’s med icinalcompounds are obtained from plants; maintain-ing as many var ieties as possible would ensurethe availability of compounds known to be use-ful, as well as new, and as yet undiscovered

compounds—e.g., the quinine drugs used in thetreatment of malaria were originally obtainedfrom the Cinchona p lant. A USDA collection of

superior germplasm established in 1940 inGuatemala was not maintained. As a conse-




quence, difficulties arose during the VietnamWar when the new an timalarial dru gs becameless effective on resistant strains of the parasiteand natural quinines were once again used.

An important d istinction exists between p re-

serving genetic resources in situ and preservinggermplasm stored in repositories. Althoughgenetic loss can occur at each location, evolu-tion will continue only in natural ecosystems.With better storage techniques, seed loss andgenetic “drift” can be kept to a m inimum. Never-theless, species extinction in situ will continue.

T he N a t iona l Ge rmplasm Sy s te m

USDA has been responsible for collecting andcataloging seed (mostly from agriculturally im-portant plants) since 1898. Yet it is important torealize that other Federal agencies also have

responsibilities for gene resource management.(See table 27.) Over the past century, over440,000 plant introductions from more than 150expeditions to centers of crop diversity havebeen cataloged.

The expeditions were needed because theUnited States is gene poor. The economically im-

Table 27.—Gene Resource Responsibilities of Federal Agencies

Type of ecosystemsunder Federal

Agency ownership/control ResponsibilitiesU.S. Department of Agriculture

Animal & Plant Health InspectionService . . . . . . . . . . . . . . . . . . . . . . . . —

Forest Service . . . . . . . . . . . . . . . . . . . . Forestlands andrangelands (U.S.National Forest)

Science & Education Administration . —

Soil Conservation Service. . . . . . . . . . . —

Controls insect and disease problems of commerciallyvaluable animals and plants.

Manages forest land and rangeland living resources forproduction.

Develops animal breeds, crop varieties, and microbial strains.Manages a system for conserving crop gene resources.

Develops plant varieties suitable for reducing soil erosion andother problems.

Department of CommerceNational Oceanic & Atmospheric

Administration. . . . . . . . . . . . . . . . . . . Oceans—between 3 Manages marine fisheries.

Department of Energy. . . . . . . . . . . . . . . .Department of Health& Human Services

National Institutes of Health . . . . . . . .

Department of the InteriorBureau of Land Management. . . . . . . .

Fish & Wildlife Service . . . . . . . . . . . . .

National Park Service . . . . . . . . . . . . . .

Department of State . . . . . . . . . . . . . . . . .

Agency for International Development. .

Environmental Protection Agency. . . . . .National Science Foundation. . . . . . . . . .

and 200 miles offthe U.S. coasts

— Develops new energy sources from biomass.

— Utilizes animals, plants, and micro-organisms in medicalresearch.

Forest lands, Manages forest, range, and desert living resources forrangelands, and production.deserts

Broad range of Manages game animals, including fish, birds, and mammals.habitats, includingoceans up to 3 milesoff U.S. coasts

Forest lands, Conserves forestland, rangeland, and desert-living resources.rangelands, anddeserts (U.S.National Parks)

— Concerned with international relations regarding gene

resources. — Assists in the development of industries in other countriesincluding their agriculture, forestry, and fisheries.

— Regulates and monitors pollution. — Provides funding for genetic stock collections and for research

related to gene resource conservation.

SOURCE: David Kapton, National Assoclatlon for Gene Resource Conservation.




portant food plants indigenous to the continen-tal United States are limited to the sunflower,cranberry, blueberry, strawberry, and pecan.The centers of genetic diversity, found mostly intropical latitudes around the world, are be-lieved to be the areas wh ere progenitors of ma-

jor crop p lants originated. Today, they containgenetic diversity that can be u sed for plant im-provement.

It is difficult to estimate the financial returnfrom the germplasm that has been collected,but its impact on the breed ing system has beensubstantial. A wild melon collected in India, forinstance, was the source of resistance to pow-dery mildew and prevented the d estruction of California melons. A seemingly u seless w heatstrain from Turkey—thin-stalked, highly sus-ceptible to red rust, and with p oor milling prop-erties—was the source of genetic resistance tostripe rust when it became a problem in thePacific Northwest. Similarly, a Peruvian speciescontributed “ripe rot” resistance to Americanpep per p lants, wh ile a Korean cucumber strainprovided high-yield production of hybrid cu-cumber seed for U.S. farmers. And a gene forresistance to Northern corn blight transferredto Corn Belt hybrids has resulted in an esti-

mated savings of 30 to 50 bushels (bu) p er acre,with a seasonal value in excess of $200 million.

20

(See tab le 28.)

The effort to store and evaluate this collected

germplasm was promoted by the AgriculturalMarketing Act of 1946, which authorized re-

gional and interregional plant introduction sta-

tions (National Seed Storage Centers) run coop-

eratively by both Federal and State Govern-

ments. The federally controlled National Seed

Storage Laboratory in Fort Collins, Colo., was

established in 1958 to provide permanent stor-age for seed. In the 1970’s, it was recognized

that the system should include clonal materialfor vegetatively propagated crops, which can-

not be stored as seed. Although their storage re-

quires more space than comparable seed stor-

ZOu. s, Dep a r tmen t of Agriculture, Agricultural Research SerV-

ice, Introduction, Class[~ication, Maintenance, Evaluation, and Docu-

mentation of Plant Germplasm, (ARS) National Research ProgramNo. 20160 (Washington, D. C., U.S. Government Printing Office,1976).

Table 28.-Estimated Economic Rates of ReturnFrom Germplasm Accessions

1. A plant introduction of wheat from Turkey was found to

2.

3.

4.

—

have resistance to all known races of common anddwarf bunts, resistance to stripe rust and flag smut,plus field resistance to powdery and snow mold. It hascontributed to many commercial varieties, with

estimated annual benefits of $50 million.The highly successful variety of short-strawed wheat,‘Gaines’ has in its lineage three plant introductions thatcontributed to the genes for the short stature and forresistance to several diseases. During the 3 years,1964-66, about 60 percent of the wheat grown in theState of Washington was with the variety ‘Gaines’. in-creased production with this variety averaged slightlyover 13 million bu or $17.5 million per year in the 3-yearperiod.Two soybean introductions from Nanking and Chinawere used for large-scale production, because they arewell-adapted to a wide range of soil conditions. All ma-jor soybean varieties now grown in t e Southern UnitedStates contain genes from one or both of these in-troductions. Farm gate value of soybean crop in theSouth exceeded $2 billion in 1974.

Two varieties of white, seedless grapes resulted fromcrosses of two plant introductions. These varietiesripen 2 weeks ahead of ‘Thompson Seedless’. Benefitsto the California grape industry estimated to be morethan $5 million annually.

SOURCE: U.S. Department of Aorlculture. Agricultural Research Service. /n-

troduct;on, Classitica~ion, Maintenance, Eveiuation, and Docu- mentation of Piant Gerrrrpiasrrr, (ARS) National Research Program No.20160 (Washington, D. C., U.S. Government Printing Of fice,1976).

age, 12 new repositories for fruit and nu t cropsas well as for other imp ortant crops, from hopsto mint, were prop osed by the N ational Germ-plasm Comm ittee as add itions to the NationalGerm plasm System (see Tech. Note 12, p. 163).(The development of tissue culture storagemethods may reduce storage costs for theseprop osed repositories.)

The National Germp lasm System is a vital link in ensuring that germplasm now existing willstill be available in the future. However, thepresent system w as challenged after the South -ern corn blight epidemic of 1970. Many scien-tists questioned whether it was large enoughand broad enough in its present form to providethe genetic resources that might be n eeded.

The devastating effects of the corn blight of 1970 actually led to the coining of the term cropvulnerability. During the epidemic, as much as15 percent of the entire yield was lost. Somefields lost their w hole crop, and entire sectionsof some Southern States lost 50 percent of their




corn. Epidemics like this one are, of course, notnew. In the 19th century, the phylloxera diseaseof grapes almost destroyed the wine industry of France, coffee rust disrupted the economy of Ceylon, and the potato famine triggered exten-sive local starvation in Ireland and mass em igra-

tion to N orth Am erica. In 1916, the red rust d e-stroyed 2 million bu of wheat in the UnitedStates and an ad ditional million in Canad a. Fur-ther epidemics of wheat rust occurred in 1935and 1953. The corn blight epidemic in theUnited States stimulated a stu dy th at led to thepu blication of a report on the “Genetic Vulner-ability of Major Crop s’’.

21It contained two cen-

tral find ings: that vu lnerability stems from ge-netic uniformity, and that some American cropsare, on this basis, highly vulnerable. (See table29.)

How ever, genetic variability, is only a hedgeagainst vulnerability. It does not guarantee thatan epidemic will be avoided . In ad dition, path-ogens from abroad can become serious prob-lems w hen they are introdu ced into new envi-ronments. As clearly stated in the study, a tri-angular relationship exists between host, path-ogen, and environment, and the coincidence of their interaction dictates the severity of disease.

z INati~nal Academy of Sciences, Genetic Vulnerability o f M@rCrops, Washington, D. C., 1972.

T he bas i s fo r ge ne t i c un i fo rmi ty

Crop uniformity results most often from soci-etal decisions on how to produce food. Thestructure of agriculture is extremely sensitive tochanges in the market. Some of the basic factorsinfluencing uniformity are:

th e consumer’s demand for high-quali ty

produce;

the food processing industry’s demand forharvest uniformity;

the farmer’s demand for the “best” variety

that offers high yields and meets the needs

of a mechanized farm system; and

the inc reased w or ld dem and fo r food ,wh ich is related to both economic and p op-ulation growth.

New varieties of crops are bred all the time,

but several can dominate agricultural produc-tion—e.g., Norman Borlaug and his colleagues inMexico pioneered the “green revolution” bydeveloping high-yielding varieties (HYV) of wheat that required less daylight to mature andpossessed stiffer straw and shorter stems. Sincethe new varieties (see Tech. Note 13, p. 163)gave excellent yields in response to applicationsof fertilizer, pesticides, and irrigation, the in-novation was subsequently introduced intocountries like India an d Pakistan. When a single

Table 29.—Acreage and Farm Value of Major U.S. Crops and Extent to WhichSmall Numbers of Varieties Dominate Crop Average (1969 figures)

ValueAcreage (millions of Total Major Acreage

Crop (millions) dollars) varieties varieties (percent)

Bean, dry . . . . . . . . . . . . . . . . . . 1.4 143 25 2 60Bean, snap. . . . . . . . . . . . . . . . . 0.3 99 70 3 76Cotton. . . . . . . . . . . . . . . . . . . . . 11.2

Coma.. . . . . . . . . . . . . . . . . . . . .

1,200 50 3 5366.3 5,200 197b 6

C

71Millet. . . . . . . . . . . . . . . . . . . . . . 2.0 ? 3 100Peanut . . . . . . . . . . . . . . . . . . . . 1.4 312 15 9 95Peas . . . . . . . . . . . . . . . . . . . . . . 0.4 80 50 2 96Potato. . . . . . . . . . . . . . . . . . . . . 1.4 616 82 4 72Rice. . . . . . . . . . . . . . . . . . . . . . . 1.8 449 14 4 65Sorghum. . . . . . . . . . . . . . . . . . . 16.8 795 ? ? ?

Soybean . . . . . . . . . . . . . . . . . . . 42.4 2,500 62 6 58

Sugar beet . . . . . . . . . . . . . . . . . 367 16 2 42Sweet potato . . . . . . . . . . . . . . . 0.13 63 48 69Wheat. . . . . . . . . . . . . . . . . . . . . 44.3 1,800 269 9 50

acorn k-iclldeg seeds, forage, and sila9e.bRelea9ed public inbreds only.

cThere were six major public lines used in breeding the major varieties of corn, so the aCtIJal number of ‘farietieS is higher.

SOURCE: National Academy of Sciences, Genetic Vu/nerabi/ity of Major Crops, Washington, D. C., 1972.




The arguments parallel those previously dis-

cussed in Congress for protection of endan-

gered species (see Tech. Note 15, p. 163). The

last decade has show n th at modes of produ ction

and development can severely affect the ecolog-

ical balance of complex ecosystems. What is not

known is how much species disruption can takeplace before the q uality of life is also affected.

4. The extent to which the new genetic technologies will affect genetic variability, germplasm storage methodologies, and crop vulnerability has not been determined.

The new genetic technologies could either in-

crease or decrease crop vulnerability. In theory,

they could be useful in developing early warn -ing systems for vulnerability by screening forinherent weaknesses in major crop resistance.How ever, the relationship betw een the genetic

characteristics of plant varieties and their pestsand pathogens is not und erstood (see Tech. Note16, p. 164).

The new technologies may also enhance theprospects of using variability, creating newsources of genetic diversity and storing geneticmaterial by:

q

q

q

q

increasing variability during cell regenera-tion,

incorporating new combinations of geneticinformation d uring cell fusion,changin g the ploidy level of plants, and

introducing foreign (nonplant) materialand dis tant ly re la ted plant mater ia l by

means of rDNA.

With the potential benefits, however, come

risks. Because genetic changes during the devel-

opment of new varieties are often cumulative,

and because superior varieties are often used

extensively, the new technologies could in-crease both the degree of genetic uniformity

and the rate at which improved varieties dis-

place indigenous crop types. Furthermore, i t

has not been determined how overcoming natu-

ral breeding barriers by cell fusion or rDNA willaffect a crop’s susceptibility to pests and dis-

eases.

5. Because pests and pathogens are constantly mutating plant resistance can be broken down, requiring the introduction of new varieties.

Historically, success and failure in breedingprograms are linked to pests and pathogensovercoming resistance. Hence, plant breederstry to keep one step ahead of mutations o r

changes in pest and pathogen populations; a

plant variety usually lasts only 5 to 15 years on

the market. There is some evidence that patho-

gens are becoming more virulent and aggres-

sive—which could increase the rate of infection,enhancing the potential for an epidemic (see

Tech. Note 17, p. 164).

6. Other economic and social pressures affect theuse of genetic resources.

The Plant Variety Protection Act has beencriticized for being a p rimary cause of p lantingun iform varieties, loss of germplasm, and con-glomerate acquisition of seed comp anies. In itsopp onents’ view, such ownership rights p rovide

a strong incentive for seed companies to en-courage farmers to buy “sup erior” varieties thatcan be p rotected, instead of ind igenous varietiesthat cannot. They also make plant breeding solucrative that the ownership of seed comp anies,is being concentrated in multinational corpora-tions—e.g., opp onents claim th at 79 percent of the U.S. patents on beans have been issued tofour companies and that almost 50 once-inde-pendent seed companies have been acquired byThe Upjohn Co., ITT, and others.

26One concern

raised about such ownership is that some of these companies also make fertilizer and

pesticides and have no incentive to breed forpest resistance or nitrogen-fixation. For theabove reasons, one public interest group hasconcluded :

27

[t]hanks to the patent laws, the bulk of theworld’s food supply is now owned and devel-oped b y a hand ful of corporations which alone,without any public input, determine whichstrains are used and how.

Num erous arguments have been advancedagainst the a bove position. Planting of a singlevariety, for instance, is claimed to be a functionof the normal desires of farmers to purchasethe best available seed, especially in the com-

2’P. R. Mooney, Seed of the Earfh (Lo n d o n : In te rn a t io n a lCoalition to r D e \ 7 el o pm e n t Action , 1979).

2T~1.1ef fo r PeoP]eS’ ~usilless ( hmm ission as Amicus C u r i a e ,

Diamond v. Chalmabarr-v, 100 S. Ct . 2204 (1980), p. 9.




petitive environment in which they operate.Moreover, hybrid varieties (such as corn), arenot covered by the plant protection laws; yetthey comprise about 90 percent of the seedtrade.

As for the loss of varieties by vegetation

displacement, statutory protection has been toorecent to counter a phenomenon that has oc-curred over a 30- to 40-year p eriod, and avail-able evidence indicates that some crops are ac-tually becoming more diverse. Since most majorfood crops are sexually produced, they haveonly been subject to protection since 1970 whenthe Plant Variety Protection Act was passed; thefirst certificates under that Act were not evenissued until 1972. Moreover, at least in the caseof wheat, as many new varieties were devel-oped in the 7 years after the pa ssage of the PlantVariety Protection Act as in the previous 17.

28

It is clear that large corporations have beenacquiring seed companies. However, the con-

20H. Rept. No. 96.1115, 96th Cong. , 2d S6?SS., p . 5 (June 20J 1980).

nection between this trend and the plant varietyprotection laws is disputed. one explanation isthat the takeovers are part of the general take-over movem ent that has involved all parts of theeconom y du ring the past decade. Since the pas-sage of the 1970 Act, the n um ber of seed com-

panies, especially soybean, wheat, and cerealgrains, has increased.29

While there were sixcompanies working with soybean breedingprior to 1970, there are 25 at th is time.

30

Thus, to date, although no conclusive connec-tion has been demonstrated between the twoplant protection laws and the loss of geneticdiversity, the use of uniform varieties, or theclaims of increasing concentration in the p lantbreeding industry; the question is stilI con-troversial and these complex problems are stillunresolved.

ZgHearin@ on H.R. 2844, sup ra note 35 (Statement of ’ Harold

Loden, Executive Director of the American Seed Trade Associa-tion).JoBrief for pharmace~t ica] Manufacturers’ Association as

Amicus Curiae, DiarnorId v. Cha)crabarty, 100 S. Ct . 2204 (1980), p .26.

S u m m a r y

The science and structure of agriculture arenot static. The technical and industrial revolu-tions and the p opu lation explosion have all con-

tributed to agricultural trends that influencethe impacts of the new technologies. Severalfactors affect U.S. agriculture in particular:

To some degree, the United States dependson germplasm from sources abroad, whichare, for the most part, located in less devel-oped countries; furthermore, the amountof germplasm from these areas that shouldbe collected h as not been d etermined.Genetic diversity in areas abroad is beinglost. The pressures of urbanization, in-dustrial development, and the demands for

more efficient, more intensive agriculturalproduction are forcing the disappearanceof biological natural resources in which thesupply of germplasm is maintained.

q This lost genetic diversity is irreplaceable.q The world’s major food crops are becoming

more vu lnerable as a result of genetic uni-

formity.The solutions—examining the risks and eval-

uating the tradeoffs—are not limited to securingand storing varieties of seed in manmaderepositories; genetic evolution—one of the keysto genetic diversity and a continuou s supp ly of new germplasm—cannot take place on storageshelves. Until specific gaps in man’s understand-ing of plant genetics are filled, and until thebreeding commu nity is able to identify, collect,and evaluate sources of genetic diversity, it isessential that natural resources providing germ-

plasm be preserved.




I s s u es an d O p t io n s — Plan t s

ISSUE: S h o u l d a n a s s e s s m e n t b e co n -

d u c t e d t o d e t e r m i n e h o w m u c h

p l a n t g e r m p l a s m n e e d s t o b e

m a i n t a i n e d ?

An understanding of how much germplasms h o u l d b e p r o t e c t e d a n d m a i n t a i n e d w o u l d

m ake the m anagem ent o f gene t i c r e sources

simpler. But no complete answers exist; nobodyknows how much diversity is being lost byvegetation d isplacement in areas mostly outsidethe United States.

OPTIONS:

A. Congress could commission a study on how much genetic variability is needed or desirable to meet present and future needs.

A comprehensive evaluation of the National

Germplasm System’s needs in collecting, eval-

uating, maintaining, and distr ibuting geneticresources for plant breeding and research could

serve as a baseline for further assessment. This

evaluation would require extensive cooperation

among the Federal, State, and private compo-

nents link ed to the N ational Germp lasm System.

B. Congress could commission a study on the need for international cooperation to manage and preserve genetic resources both in natural ecosystems and in repositories.

This investigation could include an evaluation

of the rate at which genetic diversity is being

lost from natural and agricultural systems, and

an estimate of the effects this loss will have. Un-

til such information is at hand, Congress could:

q Instruct the Department of State to have its

delegations to the United Nations Educa-tional, Scientific, and Cultural Organization

(UNESCO) and Un ited Nations Environmen -tal Program (UNEP) encourage efforts to es-

tablish biosphere reserves and other pro-

tected natural areas in less developed coun-tries, especially those within the tropical

latitudes. These reserves would serve as a

source for continued natural mutation and

variation.

q

q

q

Instruct the Agency for International De-velopmen t (AID) to place high priority on,and accelerate its activities in, assisting lessdeveloped countries to establish biosphere

reserves and other protected natural areas,providing for their protection, and supportassociate research and training.Instruct the International Bank for Recon-struction an d Developm ent (World Bank) togive high priority to providing loans tothose less developed countries that wish toestablish biosphere reserves and other pro-tected natural areas as well as to promoteactivities related to biosphere reserve pres-ervation, and the research and manage-ment of these areas and resources.Make a one-time special contribution to

UNESCO to accelerate the establishment of biosphere reserves.

Such measures for in situ preservation andmanagement are necessary for long-term main-tenance of genetic diversity. Future needs aredifficult to predict; and the resources, once lostare irreplaceable.

C. Congress could commission a study on how to develop an early warning system to recognize potential vulnerability of crops.

A followup study to the 1972 National Acad-

emy of Science’s report on major crop vul-nerability could be commissioned. Where highgenetic uniformity still exists, proposals couldbe suggested to overcome it. In addition, theavenues by which private seed companies couldbe encouraged to increase the levels of geneticdiversity could be investigated. The stud y couldalso consider to what extent the crossing of natural breeding barriers as a consequence of the new genetic technologies will increase therisks of crop v ulnerability.

ISSUE: What a re the most appropr ia te

approaches for overcoming thev a r i o u s t e c h n i c a l c o n s t r a i n t sthat limit the success of molec-ular genetics for plant improve-m e n t ?




Although genetic information has been trans-ferred by vectors and p rotoplasm fusion, DNAtransformations of commercial value have notyet been performed. Molecular engineering hasbeen impeded by the lack of vectors that cantransfer novel genetic material into plants, by

insufficient knowledge about which geneswou ld be useful for breeding pu rposes, and bya lack of understanding of the incompatibility of

c.

sis on potential pharmaceuticals derived from plants.

Establish an institute for plant molecular genetics under the Science and Education Ad- ministration at USDA that would include mul-

tidisciplinary teams to consider both basic research questions and direct applications of the technology to commercial needs and practices.

chromosomes from diverse sources. Another The discoveries of molecular plant geneticsimpediment has been the lack of researchers will be used in conjunction with traditionalfrom a va riety of disciplines. breeding programs. Therefore, each of the

three options wou ld require additional app ro-OPTIONS: priations for agricultural research. Existing

funding structures could be used for all three, A. Increase the level of funding for plant molec-

but institutional reorganization would be re-ular genetics through:1. the National Science Foundation (NSF)) and

quired for options B and C. The main argument

Z. the Competitive Grants Program of the U.S.for increasing USDA fund ing is that it is the lead

Department of Agriculture (USDA).

agency for agricultural research, for increasing

NSF and NIH funding, that they currently have B. Establish research units devoted to plant mo- the greatest expertise in m olecular techniqu es.

lecular genetics tional Institutes

T e c h n i c a l

under the auspices of the Na- Option C emp hasizes the imp ortance of the in- of Health (NIH), with empha- terdisciplinary need s of this research.

n o t e s

1. A recent example of such a mutation was th e opaque-2gene in corn, which was responsible for increasing thecorn’s content of the amino acid lysine.

Z. There is disagreement about wh at is mean t by produc-

tivity and h ow it is measu red. Statistical field data canbe expressed in various ways—e.g., output per man-hour, crop yield per unit area, or output per unit of total inputs used in p roduction. A produ ctivity meas-urem ent is a relationship amon g physical units of pro-du ction. It differs from measurements of efficiency,which relate to economic and social values.

3. Nevertheless, some parts of the world continu e to lack adequate supp lied of food. A recent study b y the Pres-idential Commission on World H unger

31estimates that

“at least one out of every eight men, women , and chil-dren on earth suffers malnu trition severe enough toshorten life, stunt p hysical growth, and d ull mentalability. ”

4. In th eory, pure lines prod uce only identical gametes,

wh ich makes th em true breeders. Successive cross-breeding will result in a mixture of gametes with vary-ing combinations of genes at a given locus on homolog-ous chromosomes.

31 RePO1.t of the pr~sidelltia] (;ornnlission on World Hung fwt ~~v~r-comin$ World Hunger: The L’hallenge Ahead, Washington, D. C.,

March 1980.

5

6

7

Normally, chromosomes are inherited in sets. Themore frequent diploid state consists of two sets in eachplant. Because chromosome pairs are homologous(have the same linear gene sequence), cells must main-

tain a degree of genetic integrity between chromosomepairs during cell division. Therefore, increases inploidy involve entire sets of chromosomes—diploid (2-set) is manipulated to triploid (3-set) or even totetraploid (4-set).The estimated theoretical limit to efficiency of photo-synthesis d uring the growth cycle is 8.7 percent. How-ever, the record U.S. State average (116 bu /acre, Il-linois, 1975) for corn, having a high photosynthetic ratein comparison to other m ajor crops, approaches only 1percent efficiency. 32

S ince a major limiting step in plant

produ ctivity lies in th is efficiency for the ph otosyn-thetic process, there is p otential for plant b reedingstrategies to imp rove the efficiency of ph otosynthesisof many other imp ortant crops. This would have a tre-mendous impact on agricultural productivity.

It is difficult to separate social values from the econom -ic structures affecting the productivity of Americanagriculture. Social pressures and decisions are complex

iZoff i~.~ o f ‘[”~~:hll[]]t)gy /l SSPSSlllPllt, 11.S. Congress, Energv for

Bioiogicai Proresses, Volume 11 : Technical Analysis (W’as hin glon ,

11.(1,: ~1.S. (;mwrnment Printing office, J u ] y 1980).



Ch. 8—The Application Of Genetics to Plants q 163

8.

9.

10.

11

and integrated—e.g., conflicts between d evelopingmaximum productivity and environmental concernsare typified by the removal of effective pesticides fromthe market. Applications of existing or new technol-ogies may be screened by th e pu blic for acceptableenvironmental impact. Conflict also exists betweenhigher productivity and higher nutritive content in

food, since selection for one often hurts the other.A critical photosynthetic enzyme (ribulose biphosphatecarboylase) is formed from information supplied bydifferent genes located independently in the chloro-plast (a plastid) and the nucleus of the cell. It is com-posed of two separate protein chains that mu st link together within the chloroplast. The larger of thesechains is coded for by a gene in the chloroplast—and itis this gene that h as been recently isolated and cloned.The smaller subu nit, however, derives from the plantnucleus itself. This cooperation between the nucleusand the chloroplast to produce the functional expres-sion of a gene is an interesting phenomenon. Because itexists, the genetics of the cell could be m anipu lated sothat cytoplasmically introduced genes can influence

nuclear gene functions. Perhaps most importantly atthis stage, plastid genes are prime candidates to clarifythe basic molecular genetic mechanisms in higherplants.The advantages to using m ass propagation techniquesfor strawberry plants are that those produced fromtissue culture are virus-free, and a plantlet produced intissue culture can produce more shoots or run ners fortransplanting.

The disad vantages are that during the first year thefruit tend s to be smaller and , therefore, less comm er-cially acceptable; the plants from tissue culture mayhave troub le adapting to soil conditions, wh ich can af-fect their vigor, especially during the first growing sea-son; and th e price per plantlet ready for planting fromtissue culture systems may be more expensive than

commercial prices for rooted shoots or runn ers boughtin bu lk .Wheat protein is d eficient in several am ino acids, in-cluding lysine. Considerable attention h as been de-voted in th e past 5 to 10 years to improvin g the nutri-tional properties of wheat. Thou sands of lines havebeen screened for high protein, with good success, andhigh lysine genes with poor success. Some high protein

varieties have been developed, but adoption by thefarmer has b een med iocre at best, partly because of redu ced yield levels. There are some “exceptions; —e.g.,the Variety “Plainsman V“ has m aintained both h ighprotein and yield levels, which indicates that there is noconsistent relationship b etween low protein and h ighyields in some varieties.

Some 42 percent of th e total land area in the tropics,consisting of 1.9 billion h ectares, contains significantforest cover. It is difficult to measure precisely theamoun t of permanen t forest cover that is being lost;however, it has been estimated that 40 percent of “closed” forest (having a continuous closed canopy) hasalready been lost, with 1 to 2 percent cleared annually.

12

13

14

15.

If the high est predicted rate of loss continu es, half of the remaining closed forest area will be lost by the year2000.

33The significance of th is loss is expressed by

Norman Myers in his report, Conversion of Tropical

Moist Forests, prepared for the Committee on ResearchPriorities in Tropical Biology of the National Academyof Science’s National Research Council: “Extrapolation

of figures from well-known group s of organisms sug-gest that there are usu ally twice as man y species in thetropics as temperate regions. If two-thirds of thetropical species occur in TMF (tropical moist forests), areasonable extrapolation from known relationships,then the species of the TMF should amount to some 40to so percent of th e plan et’s stock of species—or some-where between 2 million and 5 million species altogeth-er. In other w ords, nearly half of all sp ecies on Earthare apparently contained in a b iome that comprisesonly 6 percent of the glob e’s land surface. Probably nomore than 300,000 of these species—no more than 15percent and possibly much less—have ever been givena Latin name, and most are totally unknown . "

34

In 1975, the Committee estimated that $4 million would

be n ecessary for capital costs of each repository, withrecurring annu al expen ses of $1.4 million for salariesand operations. USDA h as allocated $1.16 million for itsshare of th e construction costs for the first facility tobe constructed at the O regon State University in Cor-vallis.

High yielding varieties (HYVs) can be defined as poten-tially high-yielding, usually semidwarf (shorter thanconventional), types that have been developed in na-tional research programs worldwide. Wheat varietieswere developed by the International Maize and WheatImprovement Center and rice varieties by interna-tional Rice Research Institute. Many improved varietiesof major crops of convention al height are not curren tlyconsidered HYV types, but they have often been incor-porated into HYV breedin g. HYVs, because of biological

a n d m a n a g e m e n t f a c t o r s , r a r e l y r e a c h t h e i r f u l lharvest potential.Although the National Germplasm System su ccessfullyhand les some 500,000 units to meet an nu al germplasmrequests, man y accessions–like the 35,000 to 40,000wheat accessions stored at the Plant Genetics andGermp lasm Institute at Beltsville, Md .–have yet to beexamined. Furthermore, the varieties released for saleby the seed companies are not presently evaluated fortheir comparative genetic differences.For comparison, the National Germplasm System func-tions on less than $10 million annually, whereas the En-dangered Species Program had a fiscal year 1980 budg-et of over $23 million. The fun ds allocated to the En-

j sRep~ r t t. th e Prmident bv a 11 .S. Interagency Task Force 011

Tropica l Forests, 7’he Worldk Tropical Forests: A Policy, S[rategv,

and Program for the C hited Slates, State Departnlent publication

No. 9117, }Vashin#on, [1.(; ., hla~~, 1980.W,V, ~lver.s, [;onktersjon Of ‘r r o ~ j c a l Moist Forests, report for the

Conmitiee on Research Priorities in Tropical Biology o f the Na-tional Research Council, National Academy of Sciences, Washing-ton, D.(:., 1980.




dangered Sp ecies Program are used for such activitiesas listing endangered sp ecies, purchasin g habitats forprotection, and law enforcement.

16. The uses of p est-resistant wh eat and corn cultivars ona large scale for both diseases an d in sects are classicsuccess stories of h ost- plant resistance. However, re-cent trends in the Great Plains Wheat Belt are disturb -

ing. The acreage of Hessian fly-resistant wheats in Kan-sas and Neb raska has decreased from ab out 66 percentin 1973 to about 42 percent in 1977. Hessian fly infesta-tions have increased wh ere susceptible cultivars havebeen p lanted. In South Dakota in 1978, in an area notnormally heavily infested, an estimated 1.25 millionacres of spring wh eat were infested resulting in lossesof $25 million to $50 million. An even greater decreasein resistant w heat acreage is expected in the n ext 2 to 5years as a result of releases of cultivars that have im -proved agronomic traits and disease resistance but thatare susceptible to the Hessian fly. Insect resistance hasnot been a significant component of commercialbreeding p rograms.

35

~sotfi{:[l ~,f ‘l’(~chno]o~v As s e s sm en t , 11. S. (Mgl’ess , Pest Manage-

rnenf Sfra~egies in Crop Protection (vol. 1, Washington, D. C.: IJ. S.(kn’ernnwnt Printing office, octoher 1979), p. 73.

17. Expressed in genetic terms, cases exist ‘(where the in -

troduction of novel sources of major gene resistanceinto commercial cultivars of crop plants has resulted inan increase in their frequency of corresponding viru-lence genes in the pathogen’ ’.” This has been reportedin Australia with wheat stem rust, barley powdery mil-dew, tomato leaf mold, and lettuce downy mildew. Evi-

dence suggests that there is considerable gen e flow inthe various pathogen populations-e.g., asexual trans-fer can quickly alter the frequ ency of virulence genes.Furthermore, pressures brought about in the evolu-tionary process have developed such a high degree of

c o m p l e x i t y i n b o t h r e s i s t a n c e a n d v i r u l e n c e m e c h -

anisms, that breeding approaches, especially those only

using single gene resistance, can be easily overcome.

3 6R , ~;. Sha t t ock , B . ~. Janss~n, R, Whitbread, and D. S. Shaw,

“h Interpretation of the Frequen cies of Host Specific Phenotypes

of Phytophthora infestans in North Wales, ” Ann. Appli. Biol. 86:249,1977.



c h a p t e r 9

A d v a n c e s i n R e p r o d u c t i v e

Bio logy and The i r E f f ec t s

o n A n i m a l I m p r o v e m e n t



Ch a p t e r 9

Page

Background . . . . . . . . . . . . . . . . . . . . . . . . . . . . 167The Scientific Era in Livestock Production . . . . . 167

Controlled Breeding. . . . . . . . . . . . . . . . . . . . . 168

Scientific Production . . . . . . . . . . . . . . . . . . . . 170

Resistance to Change . , , . . . . . . . . . . . . . . . . . 171Some Future Trends . . . . . . . . . . . . . . . . . . . . 172Technologies. . . . . . . . . . . . . . . . . . . . . . . . . . . . 173

Technologies That Are Presently Useful. . . . . . 174Sperm Storage . . . . . . . . . . . . . . . . . . . . . . . 174Artifi cial In sem in ation . . , . . . . . . . . . . . . . . 174Estrus Syn chronizatio n. . . . . . . . . . . . . . . . . 176Superovulation , . . . . . . . . . . . . . . . . . . . . . , 176Embryo Recovery . . , . . . . . . . . . . . . . . . . . . 176Embryo Transfer . . . . . . . . . . . . . . . . . . . . . 177Embryo Storage . . . . . . . . . . . . . . . . . . . . . . 177Sex Selection. . . . . . . . . . . . . . . . . . . . . . . . . 177Twinning . . . . . . . . . . . . . . . . . . . . . . . . . . . 177

Mo re Sp eculative Technolog ies . . . . . . . . . . . . 178

In Vitro Fertilization , . . , , . , , , , . . . . . . . . . 178Parthenogenesis . . . . . . . . . . . . . . . . . . . , . . 178Cloning. . . . . . . . . . . . . . . . . . . . . . . . . . . . . 178Cell Fusion . . . . . . . . . . , . . . . . . . . . . . . . . . 179Chimeras . . . . . . . . . . . . , . . . . . . . . , . . . . . 179Recomb inant DN A and Gen e Transfer. . . . . . 179

Genetics and Animal Breeding. . . . . . . . . . . . . . . 179The N ational Cooperative Dairy Herd

Im provem en t Prog ram . . . . . . . . . . . . . . . . . 180Other Species. . , . . . . . . . . . . . , . . . . . . . . . . . 181

Conclusion . . . . . . . . . . . . . . . . . . . . . . . . . . 183Impacts on Breeding . . . . . . . . . . . . . . . . , . . . 183

Dairy Cattle. . . . . . . . . . . . . . . . . . . . . . . . . . 183Beef Cattle . . . . . . . . . . . . . . . . . . . . . . . . . . 185

Page

Other Species . . . . . . . . . . . . . . . . . . . . . . . . 187Other Technologies . . . . . . . . . . . . . . . . . . . . . 188

Aquiculture . . . . . . . . . . . . . . . . . . . . . . . . . 189Poultry Breeding. . . . . . . . . . . . . . . . . . . . . .189

Issue and Options for Agriculture-Animals . . . . 1 9 0

Tables

Table No. Page

30. Heritabilit y Estimates of Some EconomicallyImportant Traits . . . . . . . . . . . . . . . . . . . . . . 169

31. Results of Su perovu lation in Farm Animals . . 174

32. Experimental Production of Identicaloffspring . . . . . . . . . . . . . . . . . . . . . . . . . . 178

33. National Co\ v-Year anc i A \ 7 er a g es for Al l

34

Official Herd Records, by Breed May 1, 1978-

Apr. 30, 1979. . . . . . , . . . . . . . . . . . . . . . . . , 181P1.e{{i(;te(j Diffc?l’enc[? of kli lk Yield of Act i~w

/\ l Blllls . . . . . , . . . . . . . . . . . . . . , . . . . . . . 181

Figures

Figure No. Page

30.31.

32.

33.

34.

Eras in LJ. S. Beef Production . . . . . . . . . . . . 168

Milk Yield/Cow and Cow Population, [ Jn i t&i

States, 1875 -1975 . . . . . . . . . . . , . . . . . . . . . . 170Milk Production per Cow in 1958-78 . . . . 17[)The Way th e Reprodu ctive TechnologiesInterrelate. , . . . . . . , . . . . . . . . . . . . . . . 175

Change in the Potential Number of Progc I l j ~per Sire From 1939 to 1979. . . . . . . . . . . . 17 6




Figure 30.— Eras in U.S. Beef Production



Ch. 9—Advances in Reproductive Biology and Their Effects on Animal Improvement q 169

Table 30.—Heritability Estimates of SomeEconomically Important Traits

Trait Heritability

Calving interval (fertility). . . . . . . . . . . . . . . . . . . . . . 10%Birth weight . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 40Weaning weight. . . . . . . . . . . . . . . . . . . . . . . . . . . . . 30Cow maternal ability . . . . . . . . . . . . . . . . . . . . . . . . . 40Feedlot gain . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 45Pasture gain . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 30Efficiency of gain. . . . . . . . . . . . . . . . . . . . . . . . . . . . 40Final feedlot weight . . . . . . . . . . . . . . . . . . . . . . . . . . 60Conformation score:

Weaning . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 25Slaughter . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 40

Carcass traits:Carcass grade . . . . . . . . . . . . . . . . . . . . . . . . . . . . 40Ribeye area. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 70Tenderness . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 60Fatthickness . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 45Retail product (percent) . . . . . . . . . . . . . . . . . . . . 30Retail product (pounds) . . . . . . . . . . . . . . . . . . . . 65

Susceptibitity to cancer eye . . . . . . . . . . . . . . . . . . . 30

SOURCE: LarryV. Cundiffand KeithE. Gregory, Beet Caft/e Breeding, USDA,

Agriculture Information Bulletin No.2B6, revised November1977, p.9.

m ay also have economic value,3but they are

much harder to m easure.

The extent to which important economic orperformance traits are genetically determinedand heritable varies from trait to trait and fromanimal to animal. (See table 30.) Heritability isdefined as the percentage of the differenceamong animals in performance traits passedfrom parent to offspring*—e.g~ bulls andheifers with su perior weight at w eaning might

average 5 pounds (lb) more than their herd-mates. Because weaning weight has an averageheritability estimate of 30 percent, the offspringof these top performing animals can be ex-pected to average 1.5 1b heavieratweaningthantheir contemporaries (0.30 X 5 = 1.5). Thisimprovement can normally be expected to bepermanent and cumulative as it is passed ontothe next generation. The improvement accumu-lates like compound interest in a savings ac-count; gains made in each generation are com-poun ded on the gains of previous generations.

3Nlirhael [. [.erner and H. P. Oonald, &foc/ern Developments in Anirr?a/ Breeding [New’ }’ork: Academic Press, 1966).

* Heri[ahility and genelir association are important in decisions

iit)oot indi~ idl ] i i l mat ings . hlost h reed ing programs are concerned~i’ it h spreading geoet ir gain rapidly throughout a population

( I l t”rd, tlork): thus two o t he r refinements for selection enter th epirt ur[’-~tvl(>riit ion irlte[’tiil, and selection d ifferential.

Like land, equipment, and cash, breeding

stock represents capital available to the com-

mercial farmer. Because all inputs must be used

efficiently, modern herd or flock managers can-

no t a f fo rd to l eave r ep roduc t ion to chance

mating in the pen or on the range. These pres-

sures for efficient production have been de-scribed as follows:4

Where dairymen are judged by the number of cows milked in an hou r, there is no place for theslow milking cow or the man who will patientlymilk her out. There is no place for the time-con-suming hurd le flock of sheep , for the small flock of chickens maintained under extensive condi-tions, or for the sow that must be watchedwhile she farrows. By degrees all classes of stock are being subjected to selection whichfavors animals that need a minimum of individ-ual attention.

The scientific basis for m odern breeding hasdeveloped slowly over the last century. Ap pliedgenetics—one part of today’s programs-hashelped modernize livestock and poultry breed-ing by elaborating on the variation of continu-ously d istributed traits in a pop ulation; carryingover what was known about rapidly reproduc-ing laboratory species, like fruit flies or mice, tothe much slower reproduction of large farmanimals; and developing the statistical tech-niques for predicting breeding values or meritand analyzing breeding programs.

s

Two examp les show the pow er of breedingtools and the increased efficiency and prod uc-tivity of today’s breeders’ stocks.

q Over the past 30 years, the average milk yield of cows in th e United States has morethan d oubled. At the same time, the num-ber of dairy cows in the Un ited States hasbeen reduced by more than 50 percent.(See figure 31.) Of this increase in outputand efficiency, more than one-fourth canbe attributed to perm anent genetic changefor at least one breed (Holsteins) partici-pating in the Dairy Herd Improvement Pro-

gram. (See figure 32.)q Poultry pr odu ction in the United States has

become the most intensive industry among

qlhid. , p. 20.~lbid. , p. 126.




Figure 31 .—Milk Yield/Cow and Cow Population,United States, 1875-1975

-

1875 ’85 ’95 ’05 ’15 ’25 ’35 ’45 ’55 ’65 1975

Year

SOURCE: J. T. Reid, “Progress in Dairy Cattle Production,” Agricultural andFood Chemistry. Past, Present, and Future, R. Teranishi (cd.)(Westport, Corm.: Avi Press, 1978).

Figure 32.—Milk Production per Cow (Holsteins) in1958=78 (New York and New England)

2-year old Holstein cows in DHIA by Al. Sires

+ 4000

+ 3000

+ 2000

+ 1000

Base1958 1962 1966 1970 1974 1978

Year

SOURCE: R. H. Foote, Department of Animal Science, Cornell University,Ithaca, N.Y. from unpublished data of R. W. Everett, CorneilUniversity.

those for farm species. For turkeys, the use

of Al in breeding for breast meat has been

so successful that commercial turkeys can

n o l o n g e r b r e e d n a t u r a l l y . T h e b i g -

breasted male, even when inclined to do so finds it physically impossible to mount the

female. As a result, a full 100 percent of thecom m erc ia l t u rkey f lock in the U ni t ed

States is replaced each year using Al. In

other species of poultry as well, productionprocesses have become equally efficient.As A. W. Nordskog has noted:

Compared with the breeding of other eco-nomically important animals, poultry breed-ing has been the first to leave the farm . . . to

become part of a sophisticated breeding in-du stry. On a commercial level, chicken s havebeen the first to be commercially exploitedby the application of inbreeding-hybridiza-tion techniques, as earlier used in corn, aswell as by methods of selective improvementusing the principles of quantitative genetics.Thus, the poultry industry, compared toother animal industries, seems to have beenthe quickest to apply modern methods of genetic improvement, including th e emp loy-ment of formally trained geneticists to handlebreeding technology plus the use of com-puters and other modern bu siness methods.

6

Sc ie n t i f i c p roduc t ion

Farm resources include land, labor, capital,

and, increasingly, new knowledge. Today, those

who innovate recapture the costs of innovating

by maintaining output while lowering costs or

by increasing output while holding costs down.

Some results of the drive toward efficiency have

included included increasing specializat ion, intensifieduse of capital and land relative to labor and in-

tegration of production p h a s e s .

P o u l t r y a n d l i v e s t o c k o p e r a t i o n s h a v e s l o w l y

become specialized over the past 50 years. Th e

farmer who used to do his own breeding, rais-

ing, feeding and slaughtering is disappearing.N o w , t h e b e e f c a t t l e i n d u s t r y i n t h e U n i t e d

States consists of: the purebred breeder who

provides breeding stock, the commercial pro-ducer, the feeder, the packer, and the retailer.

Similar specialization has occurred for most

other species—e.g., less than 15 primary breed-

ers maintain the breeding stock that produces

the 3.7 billion chickens consumed each year in

the United States. The emergence of other spe-

cialized services—such as AI providers, manage-

‘A. W. No r d s k o g , “Success and Failure of Q uantitative GeneticTheory in Poultry” in Prwceedinga of the International Conference

on Quantitative Genetics, Edward Pollacket, et al. (cd.) (Amers,

Iowa: Iowa State University Press, 1977), pp. 47-51.



Ch. 9—Advances in Reproductive Biology and The Effects on Animal lmprovement q 171

ment consultants, equipment manufacturers—has accelerated the trend toward specialization,and has given the commercial operator moretime to concentrate on his specific contributionto the chain of produ ction.

Intensification is the increasing use of some

inputs to produ ction in comparison to others.Increasing the u se of land and capital relative tolabor describes the development of U.S. agricul-ture, including livestock raising, in this century.The “factory” farm typifies this trend. Herdsand flocks are bred, . born, and raised in en-closed areas, never seeing a barnyard or theopen ran ge. The best examples of land - and cap-ital-intensive systems are those of poultry(layers, broilers, and turkeys), confined hog pro-duction, drylot dairy farming, and some vealproduction.

The greater use of land h as been encouragedby several factors, includ ing impr oved corn pro-duction for confined hog feeding, programs of preventive medicine curtailing the spread of diseases in close spaces, and environmental con-trol (light, temp erature, water, humid ity) to in-crease output under closely controlled condi-tions. However, extensive ranching for beef andsheep is still comm on in the Un ited States; thedifficulties associated with detecting estrus(“heat”) in these species and their relatively slowrates of reproduction have made it uneconom-ical to invest in them the capital necessary for

intensive farming. Furthermore, beef and sheepon extensive systems forage on marginal landthat m ight otherw ise have no u se. Beeflot feed-ing, or the fattening of cattle before slaughter ata centralized location, is the only asp ect of thebeef industry that is land-intensive; in 1977, ap-proximately one-fourth of U.S. beef cattle weref e d "

7

Linking phases of production to eliminatewaste or inefficiencies in the system has pro-gressed with great speed. For some species,such linkages now extend from breeding to the

supermarket (and, in the case of fast foodchains, to the dinner table). Integration includes

7Lyle P. Schertz , et al., Another Revolution in U.S. Farming?

USDA, ESCS, Agricultural Economic Report No. 441, December

1979 .

the linking of supply industries (feeds, medi-cines, breeding stock) with p rodu ction an d thenwith marketing services (slaughtering, dressing,packaging). Entire industries and the Govern-ment in combination have produced a complexchain of operations that makes use of Govern-

ment inspectors, the pharmaceutical industry,equipment manufacturers, the transportationindustry, and the processed feed industry in ad-dition to the traditional comm ercial farmer.

Because of this complex linkage, meat grades,cuts, and packaging have become fairly stand-ard in the American supermarket. Shoppershave come to expect these standards; consum-ers wan ting special services have learned to p aymore for them. Thus, the American farm haschanged radically over the past 30 years. Thischange has been d escribed as follows:

8

As farming enterprises grow larger, theirmanagement have to equip themselves with in-formation and resort to technologists to helpthem reach decisions and p lan for more distantgoals. Industrial developments of this kindwiden the range of farming activities, since theold style farmer, sensitive to local markets andoperating on hunches, remains as a contrast tothose for whom farming is rapidly becomingmore of a programme than a w ay of life.

Res is tance to change

New technologies in U.S. agriculture and new

ways of producing food and fiber have beenboth a cause and an effect of the movement

from farms to cities in the 20th century. Com-

mercial farmers, operating on thin or nonexist-

ent profi ts and under extreme competi t ion,have had strong reason to innovate. They have

been forced by the availability of new technol-

ogies either to do so or to watch their potential

earnings go to the neighboring farmer. Various

policies that have been adop ted to soften the im-

pa cts of the “technological treadmill ,” have

somewhat slowed the exodus from the farms.

They may have been adopted for social reasons,

but they have also become increasingly costly tosociety. The taxpayer pays for them; the con-

sumer p ays as w ell for every failure to innovateon the farms.

Wundiff , et al., op. cit., p. 9.



172 q Impacts of Applied Genetics— Micro-Organisms, Plants, and Animals

Besides a lack of capital or a lack of interest ininnovating, some farmers have resisted appliedgenetics because efficiency is not their most im-portant priority. This attitude has been de-scribed as follows:

It is easy to see why breeders are unreceptive

to the science of genetics. The business of breeding pedigree stock for sale is not just amatter of heredity, perhaps not even predomi-nantly so. The devoted grooming, feeding andfitting, the propagand a about ped igrees andwin s at fairs and show s, the dram atics of theauction ring, the trivialities of breed characters,and the good company of fellow breeders, con-stitute a vocation, not a genetic enterprise.

Farmers are tradit ionally an independent

group. Many believe that they may not directlyrecapture the benefi ts of part icipating in a

breeding program based on genetics; having no

records on one’s anim als is often p referable todiscovering proof that one’s herd is performingpoorly. On the other hand , one impact of AI hasbeen to demonstrate to farmers the value of adopting new technologies. Furthermore, theeconom ic reward of prod uction records has in-creased, since AI organizations purchase onlydairy sires with extensive records on relatives.

S o m e f u t u r e t r e n d s

Applied genetics in poultry and livestock breeding comprise a grou p of pow erful technol-ogies that have already strongly influencedprices and profits. Nevertheless, the effect of genetics is only just beginning to be felt; muchimprovem ent remains to be made in all species.It has been observed that modern genetics:

l0

. . . provides a verifiable starting point for thedevelopment of the complex breeding operationthat many populations now require . . . (which)are as far removed from simple selection as themotor car is from the bicycle.

Of these technologies, some are already inregular use, some are in the p rocess of being ap -plied, and others must await further research

and development before they become generallyavailable.

Slbid., p. 170.

‘“E. P. Cunningham, “Cu r rem Developments in the Cenetics of[,iwstock Improvement,” in 15th International Conference on Ani-

mal Blood (hwups and Biochemistry, Genelics 7:191, 1976.

Societal pressures are one of the many fac-tors that influence the introduction of thesetechnologies. Several developments around theworld will have a clear imp act on innovation ingeneral and on genetics in p articular:

An expand ing popu lation, with its growing

dem and for food p rodu cts of all kind s.The growth in income for parts of the pop-ulation, which may increase the demandfor sources of meat protein.

Increasing comp etition for the consumer’sdollar among various sources of protein,which could reduce demand for meat.

Increasing competition for prime agri-cultural land among agricultural, urban,and industrial interests. Less-than-primeland m ay also be brought back into produ c-tion as demand rises, and the same pres-sures may cause land prices to rise high .

enough to encourage greater, or intensi-fied, use of land in livestock prod uction.Increasing dem and for U.S. food and fiberproducts ‘from abroad, leading to oppor-tunities for increased profits for successfulproducers.

Changes like these will strongly affect theway Am erican farmers prod uce food and fiberproducts. The economics of efficiency and agrowing w orld pop ulation will continu e to placepressu re on the agricultural sector to innovate.In animals and animal products, efficiencies will

be found in all steps of prod uction. Efforts w illbe made to increase the number of live birthsand to reduce neonatal calf fertility, presentlyone of the costliest steps—in terms of animalslost–throughout the world. Estimates of the po-tential monetary benefits of the application of knowledge obtained from prior research in re-prod uctive physiology range as h igh as $1 bil-lion per year. Another area for great economiesin production is genetic gain. Much geneticprogress remains to be mad e in all species.

Certain technologies prom ise to increase the

ability of farmers to capitalize on the genetic im-provement of economically important traits.Suppliers of genetic material (semen, embryos)will focus increased attention on the value of their prod ucts for sale both in the United Statesand abroad.




tional dairy herd .) And economics can a lso playa role; in general, the lower an animal’s value,the less practical the investm ent in the technol-ogies, some of wh ich are relatively expen sive.

Several technologies are critical to the introductionof others.

A methodology that could rel iably induce

estrus synchronization increases the economic

feasibility of AI and embryo transfer. Likewise,

the refinement of embryo storage and other

freezing techniq ues would advance the develop-

ment of those technologies still being developed,

l ike sex selection and embryo transfer. A d -vances in in vitro fertilization will be especiallyuseful to a better understanding of basic repro-ductive processes and therefore to the devel-opment and application of the more speculativetechnologies.

The technologies interrelate.

All the technologies combined make possiblealmost total control of the reproductive processof the farm animal: a cow embryo d onor may besuperovulated and artificially inseminated withstored, frozen sperm; the embryos may be re-covered, then stored frozen or transferred di-rectly to several recipient cows whose estrouscycles have been synchronized with that of thedonor to insure continued embryonic develop-ment. Before the transfer, a few cells may betaken for id entification of male or female chro-

mosomes as a basis for sex selection. Finally,two em bryos may be transferred to each recip-ient in an effort to obta in tw ins. (See figure 33.)

Techniques not yet commercially applicable

all require embryo transfer in order to be use-

ful. They include in vitro fertilization, partheno-

genesis, production of identical twins, cloning,

cell fusion, chimeras, and rDNA technology.

The technologies described in this section are

designed to increase the reproductive efficiencyof farm animals, to improve their genetic merit,

and to enhance general knowledge of the repro-

ductive process for a variety of reasons, includ-ing concern with specific human medical prob-

lems, such as fert i l i ty regulation and better

treatments for infertility.

T e c hno logie s tha t a re p re se n t l y u se fu l

SPERM STORAGE

The sperm of most cattle can be frozen to– 196° C, stored for an indefinite period, andthen used in in vivo fertilization. Although

many of the sperm are killed during freezing,success rates [or successful conceptions (table31)] combined with other advantages of thetechnologies are enough to ensur e widespr eaduse of the technology. Short-term sperm storage(for one d ay or so) is also well-developed andwidely used.

The major advantages of storing sperm are

the increased u se of desirable sires in breeding(see figure 34), the ease of transport and spreadof desirable germplasm throughout the countryand the world, and the savings from slaughter-ing the bull after enough sperm has been col-lected. The sperm can also be tested for vene-real and other d iseases before it is used . There-fore, the use of sperm banks is expected to in-crease. Little change is anticipated in semenprocessing, other than the continued refine-ment of freezing protocols, which differ foreach species.

ARTIFICIAL INSEMINATION

The manual placement of sperm into theuteru s has played a central role in the dissemi-nation of valuable germplasm throughout theworld’s herds and flocks. Virtually all farm spe-cies can be artificially inseminated, although useof the technology varies widely for differentspecies—e.g., 100 percent of the Nation’s domes-tic turkeys are prod uced via AI compared withless than 5 percent of beef cattle. Even honey-

Table 31.—Results of Superovulation inFarm Animals

Average numberovulations normally Number of ovulations

expected with superovulation

cow . . . . . . . . . 1 6-8 Sheep. . . . . . . . 1.5 9-11

Goat. . . . . . . . . 1.5Pig . . . . . . . . . . 13 30Horse. . . . . . . . 1 1

SOURCE: George Seidel, Animal Reproduction Laboratory, Colorado State Uni-

versity, Fort Collins, Colo.




tion has been very rewarding, while selectionfor twinning in other species has not receivedmuch attention.

Twinning in nonlitter-bearing species would

greatly improve the feed conversion ratio of

producing an extra offspring. The most impor-

tant barriers, besides the high cost of embryot rans fe r t echn iques , include ext r a at t en t ion

needed for the dam during gestation, parturi-tion, and lactation.

More speculative technologies

IN VITRO FERTILIZATION

The manu al joining of egg and sp erm ou tsidethe reproductive tract has, for some species,been followed b y successful developmen t of theembryo through gestation to birth. The species

include, at this writing, the rabbit, mouse, rat,

and human. Consistent and repeatable successwith in vitro fertilization in farm species has not

yet been accomplished. The cases of reportedsuccess of in vitro fertilization, embryo reim-

plantation, and normal development in man are

beginning to be documented in the scientific

literature.

The in vitro work to date has attempted to d e-

velop a research tool so that the physiological

and biochemical events of fertilization could be

better understood. Despite the wide public at-tention it has received in the recent past, thetechnology is not perfected and will have little

II practical, commercial effect in producing in-

dividuals of any species in the near future.

Practical applications would include: a means

of assessing the fertility of ovum and sperm; a

means of overcoming female infertility by em-

bryo transfer into a recipient animal; and, when

coupled with storage and transfer, a means of

facilitating the union of specific ova and sperm

for production of individual animals with pre-

dicted characteristics.

Many of the practical applications should be-

come available within the next 10 to 20 years.Further development, along with the storage of

gametes, should allow fertilization of desired

crosses. This technology may be combined with

genetic engineering and sperm sexing in the

more distant future.

PARTHENOGENESIS

Parthenogenesis, or “virgin birth, ” is the ini-

tiation of development in the absence of sperm.

It has not been demonstrated or described for

mammalian species, and the best available infor-

mation indicates that the maintenance of par-

thenogenetic development to produce normal

offspring in mammals is p resently impossible.

CLONING

The possibility of producing genetically iden-tical individuals has fascinated both scientists

and the general public. As far as livestock are

concerned, there are several ways to obtain

genetically identical animals. The natural way is

through identical twins, although these are rare

in species other than catt le, sheep, and pri-

mates. For practical purposes, highly inbredlines of some mammals are already considered

genetically identical; first generation crosses of

these lines are also considered genetically iden-tical and do not suffer from the depressive ef-fect of inbreeding.

Laboratory method s for producing clones in-

clude dividing early embryos. The results of re-

cent experiments in the production of identical

offspring using these techniques are shown in

table 32.

Another methodology involves the insertion

of the nucleus of one cell into another, either

before or after the original genetic complementof the “receiver” cell is destroyed. Researchers

have found in cer ta in amphibia that nucleartransplantation from a body cell of an embryo

into a zygote can lead to the development of asexually mature frog.

Table 32.—Experimental Productionof Identical Offspring

Methodology Result

Dividing 2-cell embryo in 1 pair identical mouse twinshalf

Dividing morulaeain half 8 pairs of identical mouse

twins

Dividing 2-cell embryos in 5 pairs of identical sheephalf twinsDividing 4-cell embryos in 1 set identical sheepfour parts quadruplets

aAn embryo with 16 to 50 cells; resembles a mulberry.

SOURCE: Benjamin G. Brackett, School of Veterinary Medicine, University ofPennsylvania, Kennett Square, Pa.



Ch. 9—Advances in Reproductive Biology and Their Effects on Animal improvement q 181

4,000 lb of milk per lactation. In the testing year(1977-78), the su per iority su rpassed 5,000 lb p ercow. This 5,()()()-lb superiority represents 52percent more milk per lactation. The increasesin production per cow result from improvement

in both management techniques and genet ic

producing ability.

Several factors influence the rates of partic-

ipation in the NCDHIP from State to State, from

region to region, and from breed to breed. In

some States, expansion of NCDHIP membershipis not a high priority of the State Cooperative

Extension Service. In some areas, the relative

importance of dairying as an enterprise is low;

therefore, a strong local DHIA organization

does not exist. Likewise, in areas where dairy-ing is a part-time operation, dairymen have less

time and initiative for participating in the pro-

gram (although many participate in NCDHIP’sunofficial plans). Where dairymen rely on theirown b ulls and u se little AI in b reeding, progeny

testing is extremely limited. No single factor

causes dairymen in some States to take greater

advantage of the superior germplasm availableto them. The importance of strong national

leadership cannot be overemphasized in ex-

plaining the great differences among breeds in

participation rates. (See table 33.) Farsightedleadership played a large role in d eveloping th e

genetic gain of Holsteins, which represent 90percent of the U.S. dairy herd today.

The genetic gains resulting from NCDHIP are

impressive, suggesting a model for spreading

genetic superiori ty throughout the Nation’s

other herds. NCDHIP also shows the importance

Table 33.-National Cow-Year and Averages forAll Official Herd Records, by Breed

May 1, 1978-Apr. 30,1979

Cow-yearslreed (#) Milk (lb) Fat (VO) Fat (lb)

~yrshire . . . . . . . . 17,135 11,839 3.96’XO 469iuernsey . . . . . . . 57,577 10,858 4.64 504Iolstein . . . . . . . . 2,297,684 15,014 3.64 547ersey. . . . . . . . . . 89,449 10,231 4.90 501rown swiss. . . . . 24,247 12,368 4.04 500Iilking shorthorn 2,130 10,451 3.65 381Iixed and others. 83,139 13,077 3.80 497

)URCE: U.S. Department of Agriculture, Science and Education Administra-tion, Dairy Herd hnprovement Letter 55, #2, December 1979, pp. 5-6.

of combining reliable evaluation of germplasm

w i th the use o f r ep roduc t ive t echno log ies .

These technologies are of only academic in-

terest when they are used alone; i t is when

superior germplasm can be spread throughout

t h e N a t i o n t h a t t h e A m e r i c a n c o n s u m e r

benefits.

Other species

Progeny testing schemes for other species are

not as developed as they are for dairy cattle.

There are several reasons for th is lack of

testing:

q

q

q

q

Difficulty in establishing a selection objective around which to design a testing program.Milk yield and fat content were obvious

traits for selection in dairy cattle. other

species have no su ch simp le traits for selec-tion. It has been observed that, “The lack of definition of economic selection objectives

in a precise, soundly based manner is one

of the serious weaknesses of much animal

breeding of th e past. ”13

Differences in management systems. Artifi-

cial insemination is essential to the intro-

duction of superior germplasm; where it is

difficult to practice Al, elaborate testingschemes are not useful—e.g., in the Na-

tion’s beef herds, progeny testing will haveto aw a i t m ore w idespread use o f A I .

Though swine are increasingly raised inconfined housing systems, poor fertility of

boar sperm after freezing and thawing and

heat detection difficulties have limited the

use of AI.

Conflicting commercial interests. Beef bu lls,for example, continue to be sold to some

extent on the basis of fancy pedigrees and

lines, with relat ively l i t t le objective in-

formation on their genetic merit. Although

some genetic improvement programs now

exist, the beef breed associations may not

support interbreed comparisons because

some breeds would show up poorly.Conflicts between short- and long-term gains.Cross-breeding for the benefits of hybrid-

‘31.. E. A. Rowson, ‘Techniques of l.ivestoch lnlpro~renm~t ,“ Ou[-

/ook on Agricuhure 6:108, 1970.

76-565 0 - 81 - 13




ization is particularly attractive to ownersof commercial herds and flocks who con-stantly replace their stocks. This geneticimprovement is noncumulative—the im-provement d oes not continue from gener-ation to generation. At present, no strong

interest exists for impr oving th e N ation’sbeef herd as a whole, and the individualbreeder cannot effectively evaluate thegermplasm available to him.

S w ine .—There is no N ationwide testing pr o-gram for hogs in the Un ited States. * How ever, astudy of needed research prepared by theUSDA in 1976 noted th at the prod uction rate of approximately 13 pigs marketed per sow peryear in the Un ited States could be significantlyimproved. The biological potential is at least 20

to 25 p igs p er year . Similarly, a successfulbreeding program, along with other managerialchanges, could reduce the fat and increase the

lean content of pork by as much as 10 to 15 lb

per carcass.

The ARS study noted that “. . . an area that

warrants part icular at tention is the develop-

ment of a comprehensive national swine testingprogram leading to the identification, selection,

and use of genetically superior boars, togetherwith guidelines for the development and use of

sow produc t iv i ty and p ig pe r fo rm ance in -

dexes. ”14

In the case of swine, the increased use

of intensive housing, which allows reproductive

control, should increase the impetus for prog-eny testing. Likewise, pinpointing areas where

considerable improvement remains to be made

should lead to the identification of selection

objectives.

Beef.—After World War II, a few breeders

became increasingly interested in problems of

inbreeding and the economic costs of dwarfism.

By that time, some had been trained in genetics

and some breed associations and State agenciesinitiated localized testing programs for these

traits. In 1967, a “Beef Improvement Federation”

*There are several State programs—in Indiana, North Carolina,and Tennessee. Some of these programs may test only growth andnot litter size.

14[1 .s . ~epartn len~ of ~gri~ul ture , ~gricu l tu ra l Research Serv -

ire, AILS Na!ional Research Program, .Sn,ine Production, NRP No.20370,” ()(’t()hw’ 1976.

of local and breed groups was formed to try toconsolidate the different systems of the Stateimprovement programs. The Federation is now

involved in:15

. establishing uniform, accurate records,

. assisting member organizations in develop-

ing performance programs,q Encouraging cooperation among all seg-

ments of the ind ustry in u sing records,q Encouraging education by emphasizing the

use of records,

q developing confidence in performance test-

ing throughout the industry.

Despite these efforts, only about 3 percent of

beef catt le nationally are recorded. This rel-

atively low participation rate, when comparedwith NCDHIP, has both a technological and an

institutional explanation. Under the largely ex-

tensive beef raising system in the United States,AI i s d i f f icul t as long as es t rus detect ion

technologies are unavailable. Natural stud serv-

ice is usually more economical. Institution al bar-

riers also prevent the development of a strong

genet ic evaluat ion program–e.g . , the breed

associations are not al l eager to have their

breeds consistently compared with others. Like-

wise, some owners of bulls for stud service

would lose business in a strict testing scheme.

G oa ts .—Though little genetic work has been

done on goats in the past, the dairy goat in-

dustry has become more visible in the past fewyears. The desire of goat breeders to participatein NCDHIP led to the formation of a Coor-

din ating Sub -Group for Dairy Goats. A review of the research performed indicated a great need

for research in almost every area of production.As a result, AIPL developed a plan for a genetic

improvement program. The leadership in the

dairy goat industry was convinced that it could

attain genetic improvement faster and at a

lower cost via NCDHIP than it could for any

other type of research.

In 1979, AIPL received a $15,000 grant from

the Small Farms Research Funding to supperthe development of genetic evaluation proce

ISR, [,. Willhanl, ‘i(;~neti(: AN k’itv in th e 11 . S. Beef [lld Ustl’.v,

JOUIYMI Paper No. J-7923 of th e low;a Agricultur:t l i tnd Home Mnomk!s I’:xperiment Station,Ames, l ~ w i i , p r o j e c t No. 2?,()()(), n.cj .



Ch. 9—Advances in Reproductive Biology and Their Effects on Animal improvement . 183

dures for goats. Genetic evaluations for yield of dairy goat bucks w ill be available before the endof fiscal year 1980. Because limited genet ic im-provem ent for yield has occurred in dairy goatsin the p ast, these evaluations w ill probably havea significant im pact on the ind ustry. AIPL canvirtually guarantee beneficial results because of the d ata available from NCDH IP, its own exper-tise in genetics, statistics, and computer tech-nology, and the d ecades of h ighly effective re-search on genetic improvement of dairy cattlethat can be adapted for the dairy goat industry.However, funding for the goat testing programremains on a year-to-year basis.

CONCLUSION

NCDHIP has shown how important genetic in-formation is to the produ ction of meat and d airyproducts. The obstacles to such a program are

also formidable, but every failure to capitalizeon genetic potential is paid for by Americanconsumers. It has also shown that where selec-

tion objectives can b e identified and agreed on,

and where conflicting interests can be brought

together to develop a program serving all in-

terests, genetic improvement can become a cen-

tral objective in breeding programs across the

country. Without reliable, evaluative data on

breeding stock, the Nation’s breeders will havelittle interest in adopting n ew b reeding techn ol-

ogies as they become available.

Impac ts on breeding

An improvement in germplasm, like an in-

crease in the nutritional content of fertilizer or

new and improved herbicides and p esticides, in-

creases the quality of the physical capital usedon the farm. It is likely that much improvement

can still be made in the germplasm of all majorfarm animal species using existing technology.

Selecting for desired characteristics causes aspecific qualitative change; it enhances the effi-

ciency of the information contained within each

cell. The genetic information in each cell of afarm animal is either more or less desirable or

efficient than information in the cells of another

animal, depending on how it performs on im-portant traits. Superior germplasm can be usedin breeding decisions to upgrade a farmer’s

breeding or prod ucing stock. (DHIA program sare the best example of how information mightbe d istributed.)

Resources invested in genetics and in technol-ogies related to genetics will have high payoffs—

e.g.) in a classic study16

of the payoff to researchin hybrid corn and in subsequent studies of other types of genetic improvement, a highcost/ benefit ratio for such research was found .The original study also showed that the absolutemarket value of a particular product is an im-portan t factor influencing th e rate of return ona given research expenditure. In general, thegreater the aggregate value of the product, thegreater the rate of return on a research expend-iture.

17Thus, the large expenditures for meat

and animal products in the United States sug-gest a great payoff in app lied genetic research.

Beef purchases alone account for between 2 and5 percent of the Am erican consum er dollar, andthe total market value for beef is more thantwice that for corn in the United States.

DAIRY CATTLE

Total milk production has been stable forman y years. While milk prod uction p er cow hasgone steadily upward, the number of cowsdu ring the p ast 35 years has decreased p ropor-tionately. (See figure 29.) Milk production percow should continue to increase, assuming thatno radical changes in present management sys-

terns occur. The increase in production per cowcould continue even if no bulls superior to thosealready available are found , simp ly as a result of more farms switching to existing technologyand existing bulls. Moreover, bulls producedfrom this system are increasing in superiority.

The num ber of dairy cows calved as of Janu -ary 1, 1980, was 10,810)000. It has remained rel-atively stable for the past year, but may de-

Iszvi ~l.iliches, “Research [;osts and Social Returns: H.vbrid {~01’1)

and Related Innovations, “Journal of Politica/ Economy 66:419, oc-

t obe r 1958. See also R. E. Flrenson, P. E. Waggoner, an d \ ’. W. Rut-tan, “Economic Benefit From Research: An Examp le From Agricul-

tu re , ” Science 205: 1101, Sept. 14, 1979.IW. Peterson an d }’ujino Ha.vami, “Techni cal Change in Agricul-

tu re , ” Staff Papers series No. DP73-20, Depiirtment of Agrkxdture

and Applied klonomics , [ l n i tw r s i t . v of hl inmsota , St. Paul, Minn.,July 1973.




crease to around 10 million in the next d ecade if milk prod uction continues to increase.

Artif ica l Insemination. —An example of the interaction between technologies and genet-ic improvement is shown in table 34. The “pre-

dicted d ifference” (PD) in milk pr odu ction r ep-resents the ability of individual bulls to genet-ically transmit yield—the amount of milk aboveor below the genetic base that the daughters of a bull will prod uce on average due to the genesthey receive. As ind icated in table 34, the pre-dicted difference for milk yield transferred viathe bull shows an improvement from 122 to 908lb for active AI bulls in the United States overthe past 13 years.

This impressive improvement still lags behindwhat is theoretically possible. A hypotheticalbreeding program could result in an expectedyearly gain of 220 lb of milk per cow, using AI;and the biological limits to this rate of gain arenot known. In practice, the observed genetictrend in the U.S. national dairy herd is about100 lb—70 lb f rom the PDs of bu lls plus 30 lb orso from the female, most of which is actuallycarryover effect from the previous use of sup e-rior bulls.

AI organizations, many of which are coop-eratively owned by dairymen, have not rigor-ously applied the principles of AI. Their effortshave been limited by reluctance to break with

traditional selection practices, financial con-straints for pr oper testing of young bu lls to p ro-

Table 34.–Predicted Difference (PD)of Milk Yield of Active Al Bulls

Year PD milk (lb)

1967. . . . . . . . . . . . . . . . . . . . . . . 1221968. . . . . . . . . . . . . . . . . . . . . . . 1981969. . . . . . . . . . . . . . . . . . . . . . . 2051970. . . . . . . . . . . . . . . . . . . . . . . 2761971 . . . . . . . . . . . . . . . . . . . . . . . 3011972. . . . . . . . . . . . . . . . . . . . . . . 3461973. ........, . . . . . . . . . . . . . 3481974. ........, . . . . . . . . . . . . . 3381975. . . . . . . . . . . . . . . . . . . . . . . 4251976. . . . . . . . . . . . . . . . . . . . . . . 5011977. . . . . . . . . . . . . . . . . . . . . . . 5581978. . . . . . . . . . . . . . . . . . . . . . . 7481979. . . . . . . . . . . . . . . . . . . . . . . 908

SOURCE: Animal Improvement Programs Laboratory, Animal Science Institute,Beltsville Agricultural Research Center, USDA.

duce sires of cows, and too much emphasis onnonproductive traits of questionable economicvalue. The progr ess that has been mad e has re-sulted from the increased use of AI, the avail-ability of data through NCDHIP, and the actualuse of reliable genetic evaluations. If any of these three factors had been missing, far lessimprovement would have occurred,

S e m e n S t o r a g e . – It is doubtful that majortechnological changes in p rocessing semen willoccur, However, since the rate of conception isas important as the genetic merit of a sire to theeconomy of a dairy enterprise, more attentionwill be given to selecting sires of high fertility.Progress should be made in banking semen byAI studs as a hedge against costs of inflation. Inthe futu re, some of the increased costs of hous-ing and feeding bulls will probably be offset by

semen banking and earlier elimination of manybulls.

S e x e d S e m e n . —Sexing of semen to produceheifer calves (for dairymen) or bull calves (forAI organizations) has been attempted withoutsuccess for many years.

Perfect determination of the sex of progenycould practically double selection intensity intwo ways—with dams to produce bulls for test-ing in AI and d ams to p rodu ce replacements. If sexed semen is used with an AI plan, the theo-

retical improvement in milk yield would be 33lb per year, w ith 23 lb d ue to selection of dam sfor replacements.

The value of this add itional amou nt per yearmay not seem great for any individual cow, butwhen it is multiplied by a national herd of 7million cows using AI an d is accumu lated for 10years, the economic value, at $0.10/ lb, is abou t$1.1 billion—an average of$110 million per yearand $231 million d uring the l0th year. The costof sexing semen is not know n, since no one hassuccessfully done it. If a way is found, the costwou ld have to be und er $10 per breeding unitfor the procedu re to be economical.

E m b r y o T r a n s f e r .—The transfer of fer-tilized eggs from a cow to obtain progeny hasbeen accomplished with great success, Mosttransfers have involved popu lar or exotic breed-




ing animals with little regard for genetic poten-

tial.

Embryo transfer may never pay for itself in

terms of milk production of the animals pro-

duced except indirectly through bulls. Rather, itis used mostly to produce outstanding cows forsale. Other commercial applications for cattleinclude obtaining progeny from otherwise in-fertile cows, exporting embryos instead of ani-mals, and testing for recessive genetic traits,

Embryo transfer progeny must be worth

$2,500 each to justify the costs and risks. Abou t$1,500 of this represents costs due to embryot rans fe r and $1,000 the cos ts of producing

calves normally. If genetic gain from embryo

transfer comes only from dam paths, the ex-

pected gain over AI alone is 76 lb/ yr. Extra gainat $0.05/ lb above feed cost wou ld have to ac-

cumu late for 79 years before add ed gain w ouldequal even a $300 embryo transfer cost perpregnancy. If less semen is needed (allowingmore intensive bull selection), the expected gain

of 129 lb/ yr mu st accumu late for 46 years tobalance an embryo transfer cost of $300 perpregnancy.

Embryo tran sfer and perfect sexing of semenwou ld combine to impr ove genetic gain (in m ilk production) slightly. The use of less semenmight be possible throu gh ap plication of in v itrofertilization. However, feasibility based on

genetic gain would still require holding all costsdow n to arou nd $50 to $90 per conception. Thegeneral conclusion is that costs of embryo trans-fer m ust be g reatly redu ced to be economicallyfeasible if only genetic gain is considered .

E s t r u s S y n c h r o n i z a t i o n . — T h e availabilityof an effective estrus synchronization methodwould provide strong impetus for increased useof AI and embryo transfer in dairy cattle. Thedetection of estrus is an expensive operation; ef-fective control of estrus cycling also requires in-tensive management, adequate handling facil-

ities, and close cooperation between the pro-du cer, veterinarian, and AI technician.

S u m m a r y . —

Proper application of progeny testing withselection and AI can increase the genetic

gain for milk yield more than two timesfaster than is occurring today. Improvedevaluation of cows, proper economic em-ph asis on other traits, and strict adherenceto selection standards are the keys. Bio-logical limitations to this ra te of genetic im-

provement cannot be anticipated in theforeseeable future.AI of dairy cattle, with the p resent intensi-ty of sire selection, should increase the networth or p rofit of animals (increased v alueminu s extra costs of the AI program) about

$10.00/head per year. By 1990, 8 milliondairy cows in AI programs would be worthabout $800 million (8 X 10

6X 10 X 10

years) more at current market prices as aresu lt of continued use of AI.Sexing of semen when used with AI maypay for itself if the cost per breeding unit

can be kept between $10 and $20.Embryo transfer is unlikely to pay for itself genetically unless the cost is reduced to be-tween $50 and $90 per conception. How-ever, despite its high costs, it is used to pro-duce animals of exceptionally high value.(See app. II-C for an explanation of reasonsother than genetics why embryo transfer isused. )Estrus synchronization is now available foruse with heifers, and should increase theuse of AI and consequen tly the genetic im-provem ent of dairy cattle.A secondary benefit of all technologies isthe increased number of skilled personswh o can p rovide technical skills as well aseducate dairymen in all areas. Also, aunique pool of reproductive and geneticdata has been accumulated.

BEEF CATTLE

There is no single trait of overriding ire-’portance (like milk p rodu ction in dairy cows) toemphasize in the genetic improvement of beef cattle, the rate of growth is a possibility. * It isalso difficult to select for several traits at once,

q Beef and d airy cattle are usually different breeds in the LJni ted

States. In the literature and in research they are often referred toas dit”ferent species. In other countries, notably in Western European d in Japan, so-called “dual p urpose” cattle are used to produ ceboth beef and milk. In the [Jnited States, old dairy cows usually be-come hamburger.




especially when some are incompatible—e.g., itis desirable to produce large animals to sell, but

undesirable to have to feed large mothers toproduce them. There are also other complica-

tions. Growth rate has two genetic components,

for which one can select—the maternal con-

tribution (primarily milk production) and thecalf’s own growth potential. Other traits of in-

terest are efficiency of growth, carcass quality

traits (such as tenderness), calving ease, and

reproductive traits, such as conception rate tofirst service with AI,

Genetic improvement programs for beef have

two major advantages over those for dairy cat-

tle traits such as growth rate and carcass quality

can be measured in both sexes (whereas onecannot m e a s u r e the milk production of bulls);

and the trai ts are more heri table than milk production.

A r t i fi c ia l i n sem ina t ion .—Betw een 3 an d

5 percent of the U.S. beef herd is artificially in-

seminated each year. This low rate is due to sev-

eral factors, including management techniques(range v. confin ed housing), availability of re-lated technologies (especially, until recently,estrus synchronization), and the conflicting objectives of the individual breeders, ranchers,and breed a ssociations.

Because little is known about the effective-ness of Al in spread ing specific genes throu gh-out t he N ation’s beef herd s, analysts have con-

centrated on their reproductive performance.Calf losses are heavy throughout the Nation.The calf crop-the number of calves alive atweaning as a fraction of total number of femalesexposed to breeding each year—is estimated tobe between 65 and 81 percent. To put thesedata in p erspective, USDA

18has estimated that a

5-percent increase in the national calf crop

would yield a savings of $558 million per year inthe supply of U.S.-grown beef. Techniques nowavailable can produce such an increase whenthey are integrated into an ad equate manage-ment program.

The standardized measure of weaning weightin beef cattle is the weight at 205 days, adjustedfor sex of calf and age of dam. In a recent studyin West Virginia —the Allegheny Highlands Proj-ect—calf weights have averaged an increase of 10 lb per year of participation in the project, via

AI and crossbreeding. Estimates of increasedvalue of calves statewide, should the sam e testsand AI program be expanded, add up to $3.6million per year when calf prices average $50per hundredweight.

l9Rapid adoption of AI

could bring about this kind of increase in as lit-tle as 40 to 48 months.

The costs and retu rns of AI vary from farm tofarm and with the n um ber of cattle in estrus. Ingeneral, it becomes more valuable with smallerherds, more cows in estrus, higher conceptionrates, and better bulls. For purebred herds,even larger benefits have been estimated—e.g.,

in a 1969 study, the estimated increase in valueper calf when AI was used was $30.02 on pure-bred ranches compared to $3.31 on commercialranches in Wyoming.

20

A major secondary, or indirect, benefit of theuse of AI is feed saved for other uses. It hasgreatly reduced the number of sires necessaryfor stud service and, through radically im-proved milk production, the number of femalesas well. These reduced requirements togetherare equivalent to more than 1 billion bu of cornand other concentrates. This situation will be

further enhan ced as beef cattle AI expand s.synchronization of Estrus.—Differences

in the rates of application of AI between beef and dairy herds can be explained partly by thediffering management systems for the twotypes of classes of cattle. Dairy herds are kep tclose to the barn for milking and are accus-tomed to being approached by humans. In con-trast, beef herds may num ber a few thousandhead on 100,000 acres of arid pasture land. Thedetection of estrus under these conditions isdifficult.

IM[ I s, [)t,,,i,,.l ,,,(,,11 of ~~gl.i[.llltlll.p, t\gricuttural R e s e a r c h Sel~’-

i(.v, “Beet Production,” /\RS N i i t i o ! l i l ] Research Pi’Og]siIM Repel’t N o .

20360” (\ Vilshillgtoll, [).(;.: ( IS I )/ \ , ()(-t(}hel’ 19761.

MB . s, ~i ik~l . , AI . R. ~’~llsett, P. k:. Lewis, a n d F.. K . l n skq) “A

P,.(,gl.:llll RePolq 011 the Allegheny Highlands Project” Uvlorgan-

town, W. \ ’a .: West t’irginia University, January-December 19791.Z(j[). ~l . st[?l,ells and “I’. M~hl’, “Artificial In semination of Range

(htt le in \ V ~ o n li n g : An Economic Analysis, ” Wyoming Agriculturalk;xperiment S t i i t k ) n Bulletin No. 496, 1969.



Ch. 9—Advances in Reproductive Biology and Their Effects on Animal Improvement q 187

It has been predicted that the availability of prostaglindin agents for regulating estrus could

increase the number of beef calves born from

superior bulls by 10 times, and that perhaps 20

percent of th e U.S. beef cow herd could receive

at least one insemin ation artificially by 1990.21 If

this lead t o a 50-lb increase in weigh t for 10 per -

cent of the calves born, it should be worth $114million to $122 million each year, assuming 80or 85 percent net calf crop and $60 per hun-dredweight.

The implementation of recently developedestrus synchronization technology might in-crease the number of beef cows bred artificiallyby 4,000,000 in the United States. Such a pro-

gram should be successful in advancing the

calving date by one week (by decreasing thecalving interval), and in increasing the quality of

the calves produced. These new calves could be

worth about $100 million annually, less about$50 million due to extra costs associated withthe synchronization program.

S ex C o n t r o l .—S ex con t ro l w ou ld have a

dramatic effect on the beef industry. In 1971, it

was projected that by 1980 sex control couldhave an annual potential benefit of $200 mil-

lion based on 10 million female calves being re-placed by male calves produced through thesexing of semen.

22At the time of the p rediction,

the market value for steers was abou t $20 mor ethan for heifers. (Steers wean heavier and gain

more efficiently. ) Now the margin is much

greater–approximately $50. This potent ia l

method of biological control is more attractivethan the use of additives like steroids or im-

plants because of the possible hazards associ-

ated with them that preclude their use.

Em b r y o T r a n s f e r .—The possibilities forgenetic improvement in beef cattle using em-

bryo transfer have been analyzed. It appears

that embryo transfer programs can be devel-

oped to increase the rate of genetic progress for

ZIH. c). Hafs, “ potential Impact of Prostag landin on prospects fo r

F’ood f’rom Dairy Cattle,” Proc. L.ufa/vse Symposium, J. w . Lauder-da k and J. H. Sokolowski (eds. ) (Kalamazoo, Mich.: Lfpjohn, 19791,pp. 9-14.

UK . H . F’oot[? ~IMl P. Nli]ler, “ w h a t Might Sex Ratio Contro] Mean

in the Animal World,’” S.vmposium, Am. Soc. of Animal Science,

1971, pp. 1-1o.

growth rate; but the programs are much too ex-

pensive to be used over the entire population.

One problem is that the economic value of the

product of a beef cow is around 25 percent (or

even less) of that of a dairy cow. Nevertheless,

in popula t ions in which AI is used , em bryo

transfer was found to be useful for obtaining

more bulls from top cows. The females pro-duced by embryo transfer would be worth mar-

ginally more than females produced conven-

tionally, but the costs and influence of males

could spread over the population through the

use of AI. The extent of this use of embryotransfer would be very small; only a few hun-dred bulls would be p roduced per year for very

large populations, and over 99 percent of the

population would reproduce conventionally.

However, such programs could have consider-able economic benefit. Care must be taken tominimize increased inbreeding of the popula-tion w ith such a breeding scheme.

S u m m a r y . —

q

q

q

AI could substantially improve economical-

ly important traits in beef herds. However,

because of the diversity of traits consid-

ered important by different breed groups

and the lack of a national beef testing andrecording system comparable to NCDHIP,

economic estimates of its value have not

been developed.

A sexing technology to produce mostly

males (they grow faster than heifers) couldbe of enormous potent ia l benef i t to the

beef indust ry . However , no successful

technique yet exists.

Estrus cycle regulation could lead to a sub-

stantial increase in the number of beef cat-

tle in AI programs. The net benefit of thistechnology, coupled with AI, may be ashigh as $50 million p er year. Similarly, theavailability of reliable progeny recordswou ld ad d to the beneficial imp act of AI inbeef and would probably contribute sig-nificantly to its u se in beef cattle.

OTHER SPECIES

Swine.—Much progress has been made in

improving the overall biological efficiency of pork production in the United States. Improved




growth rates, feed efficiencies, carcass merit,and litter sizes have helped keep pork pricesdow n and improve its quality in the N ation’smarkets. Pork today is leaner and contains morehigh-quality protein calories than it was just afew decades ago.

AI in swine production could expand, al-thou gh it w ill be limited by the relatively poorability of swine sperm to withstand freezingand by the problem of detecting estrus. It willbe encouraged by the strong trend toward con-finemen t housing and integration of all phasesof hog production. The industry—especially theindividual, family-farm type units—would bene-fit by the establishment of a progeny testingscheme to identify superior boars. Publiclyavailable information on genetic merit woulddecrease dependence on a few corporate breed-ing organizations.

Embryo transfer in swine will be strictlylimited by difficulties in developing nonsurgicalmethods of recovery and transfer, and by thelow economic value p er animal in comparison tocattle and horses. How ever, embryo transfer isuseful in introducing new genetic material intobreeding herd s of specific pathogen-free swineand in transporting genetic material to variousregions of the world.

Sheep. —The processes of selection and of crossing specific strains, which have been so ef-fective in p oultry and hogs, have been v irtually

ignored in sheep. Selection of replacement ewesfrom the fastest growing ewe lambs born astwins and the u se of flushing to increase ovula-tion rates have led to annual increases of 1.8percent in lambing; in one test the marketweight of lambs ws increased by one lb per yearof cooperation .23

Synchronization of estrus in ewes can beachieved with prostaglandin and many differ-ent p rogestogens. The technique is u sed exten-sively in many countries, but no products forthis purpose are currently marketed in theUnited States.

AI rates abroad som etimes ap proach 100 per-cent. However, AI will not be used widely on

sheep in the United States until systems for per-formance and progeny testing are implementedthat will track the number of lambs born andtheir growth rate, and un til routine freezing of raw semen is achieved.

24

Goa ts .—The research performed on goats islargely designed for application to other ani-

mals. How ever, interest in goats in the UnitedStates and the demand for their productsthrough the w orld is increasing.

NCDHIP has just started providing sire eval-uations to goat breeders. These data, along w ithartifical insemination, should increase milk pro-du ction. The genetic data might be of particularusefulness in the less developed countrieswh ere most goat raising occurs. Greater use of all reprodu ctive technologies on v aluable Ango-ra goats m ight be expected.

Other technologies

The use of any reliable twinning or sex selec-

tion technologies will be limited until such pro-cedures can be made simple, fast, inexpensive,

and innocuous . No widespread use of these

technologies should be expected within the n extdecade.

The more esoteric techniques for manipu-

lating sex cells or the germplasm itself will haveno impact on the production of animals or

animal products within the next 20 years. Invitro manipulations, including cloning, cell fu-

sion, the production of chimeras, and the use of rDNA techniq ues, will continue to b e of intense

interest. However, it is unlikely that they will

have practical effects on farm production in the

United States in this century. Each technique

will require more research and refinement. Un-

til specific genes can be identified and located,

no direct gene manipulation will be practicable.

A polygenic basis for most traits of importance

can be expected to be the rule rather than the

exception.

Should such techniques become available,

limited use for produ cing b reeding stock can b eexpected. Experience with early users of AI and

z3K;. K . I l l skeep, personal Lmllllllllllicatioll, 1980. Z41bid.




embryo tran sfer is strong evidence for the p re-dicted use of the technologies, no matter whattheir economic justification. (See app. II-C.)

A major, secondary effect of animal researchin reproductive biology is increased under-

standing leading to the possible solution of human problems—e.g., the concept, efficacy,and safety of the original contraceptive pill wasdeveloped and established in animals. It in-volves the sam e pr inciple as estrou s cycle reg-ulation discussed above.

AQUACULTURE

Aquiculture is the cultivation of freshwaterand marine species (the latter is often referred

to as mariculture). While fish culture is about6,000 years old, scientific understanding of itsbasic principles is far behind that of agriculture.Aquicultur e is slowly being transformed into amodern multidisciplinary technology, especiallyin the industrialized countries. Increasingawareness of human nutritional needs, over-fishing of natural commercial fisheries, and ris-ing worldwide d emand for fish and fish p rod-ucts are trends that indicate a growth in inter-est in aquiculture as a means to meet the foodneeds of the world’s population.

As part of the trend toward the high tech-nology and dense culturing of intensive aqua-culture systems in the industrialized countries,p rob lem s o f r ep roduc t ive con t ro l , ha t che ry

technology, feeds technology, disease control,and systems engineering are all being investi-

gated. Reproductive control and genetic selec-t ion are important because most commercial

aquicul ture operat ions must now depend on

wild seedstocks. Very little information on the

animals in culture is available.

With all three of the aquiculture genera (fish,

mollusks, and crustaceans), selective breeding

programs have long been established, healthy

gene pools are available, and advantageous hy-

bridizations have been developed. In fish rais-

ing, culture systems often demand sterile hy-brids, especially of carp and tilapia, Selective

breeding of salmon h as been limited b y politicalpressures. Very little work has been conducted

with catfish, the largest aquiculture industry in

the United States. The use of frozen sperm,

which has been successful, should increase be-

cause of the savings in transport costs. Although

culture systems for mollusks are fair ly well-

defined, little applied genetics work has been

done with these popular marine species. Some

success has been r epor t ed in se l ec t ion fo r

g row th r a t e and d i sease r e s i s t ance o f t heAmerican oyster, and selection for growth rate

of the slow-growing abalone is underway. The

crustaceans, of which the Louisiana crayfish is

the largest and most viable industry, are the

least understood. Successful hybrids of lobsters

have been developed.

Aquaculture suffers from an insufficient re-search base on the species of interest. However,growing appreciation of and demand for ma-rine species should result in increased supportfor basic and d evelopm ental work on a ll aspects

of control, including basic reproductive biology.POULTRY BREEDING

The quantitative breeding practices of com-mercial breeders have changed very little over

the last 30 years. Highly heritable traits, such

as growth rate, body conformation, and egg

weight, are perpetuated by mass selection be-

cause l i t t le advantage is gained from hybrid

vigor. Low heritable traits (egg production, fer-

tility, and disease resistance) are perpetuated by

crossbreeding and identified through progenyand family testing.

The goals of the industry are to increase eggproduction of the layers–both in quality and

quantity–and, with broilers and turkeys, to im-

prove growth rate, feed efficiency, and yield, as

well as to reduce body fat and the incidence of

defects.

The technologies of AI and semen preser-

va t ion have acce le ra t ed the advances m adethrough quantitative breeding technology. AI is

widely used in commercial turkey breeding be-cause of the inability of modern strains to mate.It makes breed ing tests m ore efficient, steps up

selection p ressure on the m ale line, redu ces thenumber of necessary breeder males, and in-creases the number of females that may be

mated to one male. Semen diluents were intro-

duced to the turkey industry about 10 years agoto lower the cost of AI. Currently, a little over




half of the turkeys are inseminated with diluted

semen.

Preservation of poultry semen by freezing is

now practiced by several primary breeders. Al-though freezing chicken semen causes it to losesome potency, the practice allows increased ge-

netic advancement and the distribution of ge-netic material worldwide.

The amount of genetic variation available forbreeding stock is not expected to dim inish in thenear future. Ceilings fo r certain traits will even-tually be reached, but certainly not in the1980’s. Advances in breeding laying chickenswill be less dram atic than in the p ast, but effortswill continue to d evelop new genetic lines andto improve reserve lines and crosses to meet fu-ture needs.

Th e growth rate of broilers will continue t .

increase at 4 percent a year, which suggeststhat bird s w ill be reaching 4.4 lb in 5 weeks bythe 1990’s. Breeding for stress resistance will beincreasingly imp ortant, not only because of theincreased use of intensive produ ction systems,but also to meet the physiological stresses re-sulting from faster growth and greater weight.

AI w ill assum e increasing importan ce. Recentadvances in procedures for long-term freezingof chicken semen will allow breeders to extendthe u se of outstand ing sires. The sale of frozenseman may eventually substi tute, in part , for

the sale of breeder males.

Dwarf broiler breeders will also assume in-creasing importance over the new few years.The dw arf breeder female is app roximately 25-percent smaller than the standard female, andeven thou gh the d warf’s egg is smaller and the

progeny’s growth rate slightly less than that of the standard broiler, the lower cost of produc-ing broiler chicks from the dw arf breeder m orethan offsets the slight loss in their grow th rate.Dwarf layers and the dw arf breeder hen s couldreduce production costs by 20 percent and 2

percent, respectively.

There is some interest among poultry breed-

ers in cloning, gene transfer, and sex control

but progress toward successful technologies is

slow.

I s sue and Opt ions fo r Agr icu l tu re—Animals

I SSU E : Sh o u ld t h e U n i t ed S t a t e s i n -

c r eas e s u p p o r t f o r p r o g r ams ina p p l ie d g e n e t i c s fo r a n i m a l s a n da n i m a l p r o d u c t s ?

Advocates of a strong governmental role in

support of agricultural research and develop-ment (R&D) have traditionally referred to thesmall size of the prod uction u nit: U.S. farms aretoo small to sup port R&D activities. Throu ghou tthis century a complicated and extensive net-work of Federal, State, and local agriculturalsupport agencies has been developed to assistthe farmer in app lying the new knowledge pro-

duced by research institutions. This private/ public sector cooperative network has pro-duced an abundant supply of food and fiber,sometim es in excess of domestic dem and . Social-ly oriented p olicies have been adop ted to soften

the impacts of new technology and to rescue the

marginally efficient farmer from bankruptcy.Curr ent projections of U.S. and w orld p opu la-

tion growth show increasing demand for allfood products. Other predictable trends withimplications for agricultural R&D, include:

q

q

q

q

q

growth in income for some populations,which will probably increase the demandfor sources of meat protein;increasing com petition am ong varioussources of protein for the consumer’sdollar;increasing awareness of nutrition issues

among U.S. consumers;increasing competition for prime agricul-tural land among agricultural, urban, andindustrial interests;increasing d emand for U.S. food an d fiber




products from abroad, leading to oppor-

tunities for increased profits for successful

producers; and. increasing demands on agricultural prod-

ucts for produ ction of energy.

OPTIONS:

A. Governmental participation in, and funding of, programs like the National Cooperative Dairy Herd Improvement Program (NCDHIP) could be increased. The efforts of the Beef Cattle Improvement Federation to standardize procedures could be actively supported, and a similar information system for swine could beestablished.

The fastest, least expensive way to upgradebreeding stock in the United States is through

effective use of information. Computer technol-ogy, along with a network of local represent-atives for d ata collecting, can p rovide th e indi-vidual farmer or breeder with accurate infor-mation on the germplasm available, so that hecan then make his own breeding decisions. Inthis way, the Nation can take advantage of pop-ulation genetics and information han dling capa-bilities to upgrade one of its most importantforms of capital: poultry and livestock. Breedassociations and large ranchers who sell thesemen from their prize bulls based on ped igreesrather than on genetic merit may act as barriers

to the effectiveness of such an objective infor-mation system.

The benefits of such pr ograms w ould accrueboth to U.S. consumers, in reduced real pricesof meat and animal produ cts, and to prod ucerswho participate in the programs, in increasedefficiency of production. Consumers spend sucha large part of their incomes on red meat thatevery increase in efficiency represents millionsof dollars saved. Beef producers too, shouldwelcome any assistance in upgrading theirstocks. The price of semen has remained rel-

atively stable, and semen from bulls ratedhighly on certain economic traits costs only afew d ollars more than that from average bu lls.

How ever, efficiency of prod uction is not th eonly value to be u ph eld in U.S. agriculture—e.g.,in milk pr odu ction complex policies have been

designed to maintain constant milk supplieswithout large fluctuations in price.

The NCDHIP model program for dairy cattlehas shown that an effective national programrequires the participation by the varied in-

terests in p rogram policymaking in an extensionnetwork, for local collection and validation of data an d for education and of expertise in datahandling and analysis. Also important is astrong leadership role in establishing the pro-gram. This option implies that the Federal Gov-ernment would play such a role in new pro-grams and expand its role in existing ones.

B. Federal funding of basic research in total animal improvement could be increased.

The option, in contrast with option A,assumes that it is necessar y to maintain or ex-

pand basic R&D to generate new knowledgethat can be applied to the production of im-proved animals and animal products.

Information presented in this report supportsthe conclusion that long-term basic research onthe physiological and biochemical events inanimal d evelopm ent results in increasing th e ef-ficiency of animal production, both in totalanimal numbers and in quality of product. In-creased un derstanding of the interrelationshipsamong various systems—including reproduc-tion, nutrition, and genetics—gradu ally leads to

the development of superior animals that effi-ciently consume food not palatable to humansand are resistant to d isease.

Earlier studies also support the importance of basic research–e.g., the National ResearchCouncil found in 1977 that”. , . not as much fun-damental research on animal problems hasbeen conducted in recent years . . . it shouldreceive increased fun ding. ”

25USDA also found,

in a review of various conference proceedings,congressional hearings, special studies, andother p ublished m aterials on agricultural R&Dpriorities, strong su pp ort for more research onthe basic processes that contribute to rep roduc-tion an d p erforman ce traits in farm animals:

2sN~~t iollill [~f,$(,ill.(,ll (hllll(;il, Worki FIIod aIId Nutr i t ion .stlJd.Vt

‘/’/If; Polfmlia/ (,’onlrilmlions qf’ Rfksfwrch ILVilShill#Oll, l). (:. i l l l t t l o l ’ ,

1977 ) , p. 9 7 .




Specific livestock research areas identified ashaving significant potential for increased p ro-duction both in the Un ited States and d evelop-ing countries include: 1) control of reprodu ctiveand respiratory diseases, 2) developing geneti-cally superior animals, 3) improving nutrition

efficiency, and 4) increasing the reprod uctiveperformance of all farm animal species.

26

z611. S. Depa r tmen t of A#ictdture, Science an d Education Ad-

ministration, Agricuhura/ and Food Research Issues an d Priorities

(Washington, D. C.: author, 1978), p. xiii.

Regardless of the effectiveness of presentpopulation control programs or of currenttrend s in individu al decisions abou t family size,the output of the Nation’s agricultural activitiesmust increase over the next decades if sufficientfood is to be available for the world’s popula-tion. Basic research is the source from whichnew app lications to increase p rodu ctivity arise.



P a r t III

I n s t i t u t i o n s a n d Soc ie ty

Chapter 10. The Question of Risk . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 197

Chapter 11. Regulation of Genetic Engineering. . . . . . . . . . . . . . . . . , . . . . . . . . . . . . . .211

Chapter 12. Patenting Living Organisms. . . . . . . . . . . . . . . . . . . . . . . , . . . . . . . . . . . . .237

Chapter 13. Genetics and Society . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .257



C h a p t e r 10

Th e Qu es t io n o f R i sk



C h a p t e r 10

Page

Introduction. . . . . . . . . . . . . . . . . . . . . . . . . . . . 197The Initial Fear of Harm . . . . . . . . . . . . . . . . . . . 197

Classif ication o f Poten tial H arm . . . . . . . . . . . . 198Iden tification o f Possib le H arm . . . . . . ., . . . . zooEstimates of Harm: Risk. . . . . . . . . . . . . . . . . .200

The Status of the Current Assessment of Physical Risk . . . . . . . . . . . . . . . . . . . . . . . 201

Perception of Risk . . . . . . . . . . . . . . . . . . . . . .203Burden of Proof . . . . . . . . . . . . . . . . . . . . .. 203

Other Concerns . . . . . . . . . . . . . . . . .........204Concerns Raised by Industrial Applications .. .204Concerns Raised b y the Imp lications of the

Recombinan t DN A Controversy for Gen eralMicrobiology. . . . . . . . . . . . . . . . . . . . . .. ..204

Concerns Raised b y the Imp lications of theRecombinan t DNA for Other Gen eticManipulation . . . . . . . . . . . . . . . . . . . . .. ..206

Page

Ethical an d Moral C on cerns . , . . . . . . . . . . . . .207Conclusion . . . . . . . . . . . . . . . . . . . . .........207

Figures

Figure No. Page

35. Flow Chart of Possible Consequ ences of UsingGenetically Engineered Micro-Organisms , . . 199

36. Flow Chart to Establish Probability of HarmCaused by the Escape of a Micro-OrganismCarrying Recombinant DNA . .............201

37. Alternative Method s for Transferring DN A FromOne Cell to Another. . . . . . . . . . . . . . . . . . . . 206



Ch ap te r 10

The Q ues t i on o f R i s k


The perception that the genetic manipulationof micro-organisms might give rise to unfore-

seen risks is not new. The originators of chem-

ical mutagenesis in the 1940’s were warned that

harmful uncontrolled mutations might be in-

duced by their techniques. In a let ter to theRecombinant DNA Advisory Committee (RAC) of

the Nation al Institutes of Health (NIH) in Decem-

ber of 1979, a pioneer in genetic transformation

at the Rockefeller University, wrote: “. . . I did

in 1950, after some deliberation, perform the

first drug resistance DNA transformations, and

in 1964 and 1965 took part in early warnings

against indiscriminate ‘ transformations’ thatwere then b eing imagined. ”

1

Yet none of this earlier public concern led toas great a controversy as has research with re-

combinant DNA (rDNA). No doubt it was en-

couraged because scientists themselves raised

questions of potential hazard. The subsequent

open debates among the scientists strengthened

the public’s perception that there was legitimate

cause for concern, This has led to a continuing

attempt to define the potential hazards and the

chances that they might occur.

‘ Rollin 1). Ho[chkiss, Recombinant DNA Research, vol. 5, N IH pub-lication No. 80-2130, March 1980, p . 484.

T h e i n i t i a l f e a r o f h a r m

For the purposes of this discussion, harm (or

injury) is defined as any undesirable conse-

quence of an act. Such a broad definition is war-ranted by the broad targets for hypothetical

harm that genetic manipulation presents: injuryto an individual’s health, to animals, to the en-

vironment.

The inital concern involved injury to human

health. Specifically, it was feared that combin-

ing th e DN A of simian virus 40, or SV40, with an

Escherichia coli plasmid w ould establish a n ewroute for the dissemination of the virus. Al-

though the SV40 is harmless to the monkeys

from which it is obtained, it can cause cancer

when injected into mice and hamsters. And

while it has not been shown to cause cancer in

humans, it does cause human cells to behavelike cancer cells when they are grown in tissue

culture. What effect such viruses might have if

they were inserted into E. coli, a normal in-habitant of the human intestine, was unknown.This uncertainty, combined with an intuitive

judgment, led to a concern that somethingmight go w rong. The d angerous scenario wentas follows:

q SV40 causes cells in tissue culture to be-have like cancer cells,q SV40-carrying E. coli might be injected ac-

cidently into hum ans,q humans would be exposed to SV40 in their

intestines, andq an epid emic of cancer wou ld result.

This chain of connections, while loose, wasstrong enough to raise questions in at least somepeople’s mind s.

The virus SV40 has never actually beenshown to cause cancer in humans; but the po-

tential hazards led the Comm ittee on Recombi-nant DNA Molecules of the National Academy of Sciences (NAS) to call in 1974 for a deferment of any experiments that attemp ted to join the DNAof a cancer-causing or other animal virus to vec-tor DNA. At the same time, other experiments,

197

76-565 0 - 81 - 14




that w ere thought to h ave a potential for harm–particularly those that were designed totransfer genes for potent toxins or for resist-ance to antibiotics into bacteria of a differentspecies—were also deferred. Finally, one othertype of experiment, in wh ich gen es from higher

organisms might have been combined with vec-tors, was to be postp oned . The fear was that la-tent “cancer-causing genes” might be inadver-tently passed on to E. coli.

Throughout the moratorium, one point wascertain: no evidence existed to sh ow that harmwould come from these experiments. But it wasa p ossibility, The scientists wh o originally raisedquestions w rote in 1975: “. . . few, if any, believethat this methodology is free from risk. ”

2It was

recognized at that t ime that “. . . estimating therisks will be difficult and intuitive at first butthis will improve as we acquire additionalknowledge. ” 3 Hence two principles were to befollowed : containm ent of the micro-organisms

(see table 35, p. 213) was to be an essential p artof any experiment; and the level of containmentwas to match the estimated risk. These prin-ciples were incorporated into the Guidelines forResearch Involving Recombinant DNA Mole-cules, promulgated by NIH in 1976.

But th e original fears surrou nd ing rDNA r e-search progressed beyond concern that humansmight be harm ed. Ecological harm to plants, ani-mals, and the inanimate w orld were also consid-

ered. And other critics noted the possibility of moral and ethical harm, which might disruptboth society’s structure and its system of values.

C l a s s i f i c a t i o n o f p o t e n t i a l “

ph y sic a l h arm

Some combinations of DNA may be harm fulto man or his environment—e.g., if an entireDNA copy of the p oliovirus genetic material iscombined w ith E. coli plasmid DN A, few w ouldargue against the need for careful handling of this material.

For practical purposes, the potential harmassociated with various micro-organisms is

Viecombimm[ DNA Research, vol. 1, DHEW pub lication No. (NIH)76-1138, Augus t 1976, p. 59.31bid.

shown in figure 35. Each letter (A through L)represents the consequence of a particular com-bination of events and micro-organisms. For ex-ample, the letters:

A)C

B,D

E, I

F)J

H,L

G)K

represent the intentional release of micro- organisms known to be harmful to the

environment or to man—e.g., in biologi-cal warfare or terrorism.represent the inadvertent release of

micro-organisms known to be harmful toth e environment or to man—e.g., in acci-

den t s a t h igh-con ta inm ent f ac i l i t i e s

where work is being carried out with

dangerous micro-organisms.

represent the intentional release of micro- organisms thought to be safe but which prove harmful—when the safety of orga-nisms has been misjudged.

represent the intentional release of micro- organisms which prove safe as expected—e.g., in oil recovery, mining , agricultur e,and pollution control.represent the inadvertent release of micro-organisms which have no harmful consequences— e.g., in ordinary accidentswith harmless micro-organisms.represent the inadvertent release of

micro-organisms thought to be safe bu twhich prove harmful—the most unlikelypossible consequence, because both anaccident m ust occur and a misjud gment

about the safety mu st have been mad e.Discussions of physical harm have recognized

the possibility of intentional misuse but haveminimized its likelihood. The Conv ention on theProhibition of the Development, Production,and Stockpiling of Bacteriological (Biological)and Toxin Weapons and on their Destructionwhich was ratified by both the Senate and thePresident in 1975, * states that the signatorieswill “never d evelop . . . biological agents or tox-ins . . . that h ave no justification for pr oph ylac-tic, p rotective, or other peaceful purp oses. ”Such a provision clearly includes micro-orga-nisms carrying rDNA molecules or the toxins

4[;onvention of the Prohibition of the Developmen t, Production,and Stockpiling of Bacleriolo~ical (Biological) an d ‘1’oxin Weaponsand on Their Destruction, Washin@on, Umdon , and Moscow,Apr. 10, 1972; entered into fo rce on M ar. 26, 1975 (26 [J. S.”r., 583).

*As of 1980, 80 countries ha te ratified the treaty; another 40have signed but n ot ratified.



Ch. 10—The Question of Risk . 199

Figure 35.—Flow Chart of Possible Consequences of Using Genetically Engineered Micro-Organisms


produced by them. It must be assumed thatthose who signed did so in good faith.

While there is no way to judge the likelihoodof developments in this area, the problems thatwould accompany any attempt to use pathogen-ic micro-organisms in warfare—difficulties incontrolling spread, protection of one’s owntroops and population —tend to discourage theuse of genetic engineering for this purpose. *Similarly, the danger that these techniques

might be used by terrorists is lessened by thescientific sophistication needed to construct a

more virulent organism than those that can

* A 11 hou~h storkpil in ~ of ” biological ma rf’are agents is prohibited,l’(’S(’ill’(’tl into 11(W’ il#lllS is Ilot.

1

already be obtained —e.g., encephalitis virusesor toxin-producing bacteria like C. botulinum orC. tetani.

Some discussions have centered around thepossibility of accidents caused by a break in con-tainment. Construction of potentially harmfulmicro-organisms will probably continue to beprohibited by the Guidelines; exceptions will bemade only under the most extraordinary cir-cumstances. To date, no organ ism know n to be

more harmful than the organism serving as thesource of DNA has been constructed.

However, the biggest controversy has cen-tered around unforeseen harm—that micro-organisms thought safe might prove harmful.




Discussion of this kind of harm is hindered by

the difficulty not only of quantifying the prob-ability of an occurrence but also of predicting

the type of damage that might occur. The differ-

ent types of damage that can be conjured u p arelimited only by imagination. The scenarios have

included epidemics of cancer, the spread of oil-

eating bacteria, the uncontrolled proliferation

of new plant life; and infection with hormone-

producing bacteria.

The risk of harm refers to th e chance of harmactually occurring. In the present controversy,it has been d ifficult to d istingu ish the p ossiblefrom the probable. It is, for instance, possiblethat an individual w ill be killed by a meteor fall-ing to the groun d, but it is not probable. Analog-ous situations exist in genetic engineering. It isin this analysis that debate over genetic engi-neering has some special elements: the uncer-

tainty of wh at kind of harm could occur, the u n-certainty about the magnitude of risk, and theproblem of the perception of risk.

Ident i f ica t ion o f poss ib le harm

The first step in estimating r isk is identifyingthe poten tial harm. It is not very mean ingful toask: How much risk does rDNA pose? The con-cept of risk takes on mean ing only when ha rm isidentified. The question shou ld be: What is thelikelihood that rDNA will cause a specific dis-ease such as in a single individual or in an entirepop ulation? The magnitud e of the possible harmis incorporated in the qu estion of risk, but d if-fers in the two cases. A statement about the risk of death to one person is different than oneabout the risk of death to a thousand. The rightquestions must be asked about a specific harm.

Since no dangerous accidents are known tohave occurred, their types remain conjectural.Identifying potential harm rests on intuition an dargum ents based on an alogy. Even a so-calledrisk experiment is an approximation of subse-quent genetic manipulations. That is why ex-perts disagree. No incontestable “scientificmethod” dictates which analogy is useful or ac-ceptable. By their very nature, all analogiesshare some characteristics with the event underconsideration but differ in others. The goal is to

discover the one that is most similar and toobserve it often. This process then forms thebasis for extrapolation.

For examp le, it has been argu ed th at ecologi-cal dam age can be caused by the introdu ction of plants, animals, and m icro-organisms into new

environm ents. Scores of examples from historysupp ort this conclusion. The introduction to theUnited States of the Brazilian water hyacinth inthe late 19th century has led to an infestation of the Southern waterways. Uncontrolled spreadof English sparrows originally imported to con-trol insects has made eradication programs nec-essary. Countless other examp les are confirma-tion that biological organisms may, at times,cause ecological damage w hen introdu ced into anew environment. Yet there is no agreement onwhether such analogies are particularly rele-vant to assessing potential dan gers from genet-

ically engineered organisms. It could be ar-gued-e.g., that a genetically engineered orga-nism (carrying less than 1 percent new genes) isstill over 99 percent the same as the original,and is therefore not analogous to the “totallynew” organism introduced into an ecosystem.Some experts emphasize the differences be-tween the situations; others emp hasize the simi-larities.

Other analogies have been raised. Newstrains of influenza viru s arise regu larly. Somecan cause epidemics because the population,

never before exposed to them, carries no pro-tective antibodies. Yet can this analogy suggestthat r elatively harm less strains of E. coli mightbe transformed into epidemic pathogens? Thereis disagreement, and debates continue aboutwhat “could happen” or what is even logicallypossible.

Es t imates o f harm: r isk

Assuming that agreement has been reachedon the possibility of a specific harm, what can bedone to ascertain the probability? What is the

likelihood that d amage w ill occur?Damage invariably occurs as the result of a

series of events, each of wh ich has its own p ar-ticular chance of occurring. Flow charts havebeen prep ared to id entify these steps. A typical




analysis determines a probability value for eachstep-e. g., in figure 36 step II the probability of

escape can be estimated based on the historical

record of experiments with micro- organisms.

Depending on the degree of containment, the

probability varies. I t i s a lmost cer ta in that

experiments on an open bench top, using noprecautions, will result in some escape to the

surrounding environment—a much less likely

event in maximum containment facilities. (Seetable 35.)

Two p oints should be noted. First, each p rob-ability can be minimized by appropriate controlmeasu res. Second , the pr obability that the finalevent w ill occur is equal to or less likely than theleast likely link in the chain. Because the prob-abilities must be multiplied together, if theprobability of any single step is zero, the p rob-

ability of the final outcome is zero; the chain of events is broken.

THE STATUS OF THE CURRENT ASSESSMENTOF PHYSICAL RISK

A successful risk assessment should provide

information about the likelihood and magnitude

of damage that might occur under given cir-cumstances. It is clear that the more types of dam age that are identified, the more risk assess-ments must be carried out.

Figure 36.— Flow Chart to Establish Probability of

Harm Caused by the Escape of a Micro-OrganismCarrying Recombinant DNA

Event Probability

L Inadvertent incorporation of hazardous gene P I

into micro-organism

11. Escape of micro-organism into environment P,

Ill. Multiplication of micro-organism and P3

establishment in ecological niche

V. Production of factor to cause disease

P*

NOTE: P, will always be smaller than any of the other probabilities.


Although the original charter of RAC under-scored the importance of a risk assessment pro-gram, it w as not u ntil 1979 that th e details of aformal program were published. For 5 years,risks were assessed on a case-by-case basisthrou gh: 1) experiments carried ou t un der con-

tract from NIH, 2) experiments that were de-signed for other purp oses but w hich p roved tobe relevant to the question of risk, and 3) con-ferences at wh ich findings w ere examined.

From the start, it was difficult to design ex-periments that could supply meaningful infor-mation-e.g., how does one test the possibilitythat “massive ecological disruptions mightoccur?” Or th at a new bacterium with h armfulunforseen characteristics will emerge? Stillsome experiments were proposed. But becausethese experiments had to be approximations of

the actual situation, the applicability of theirfindings was debated. Here too, experts couldand did disagree—not about the findings them-selves, but abou t their interpretation.

For example, in an imp ortant experiment d e-signed to test a “w orst case situation, ” a tum orvirus called polyoma was found to cause no

tum ors in test animals when incorporated intoE. coli.

5* Since just a few molecules of the viral

DNA are known to cause tumors when injecteddirectly into animals, it was concluded thattumor viruses are noninfectious to animalswhen incorporated into E. coli. If polyoma virus,which is the most infective tumor virus knownfor hamsters, cannot cause tumors in the rDNAstate in E. coli, it is unlikely that other tumorviruses will do so. This conclusion has had wide-spread, but not unanimous, acceptance. It hasbeen argued that there might be “somethingspecial” about polyoma that prevents it fromcausing tumors in this altered state; othertum or viruses might still be able to d o so. At onemeeting of RAC, in fact, it w as suggested thatexperiments with several other viruses be car-ried out to confirm the generality of the finding.

But how man y more viruses? What is enough?

‘M . A. Israel, H. W. Chan, W. P. Rowe, and M . A. Martin, “Molec-ular Cloning of Po lyoma Virus DNA in Eacherichki Cof i : PlasmidVector Systems,” Science 203:883-887, 1979.

*Some combinations of free plasmid and tumor virus DN A didcause infections.




the conclusion is that normal hormone levels

would change by less than 10 percent. Similar

cond i t ions fo r i n t e r f e ron p roduc t ion cou ld

release approximately 70ug or the maximum

daily dose currently used in cancer therapy.Long-term effects of such exposure are current-

ly unkn own; therefore, experiments u sing h igh-producing strains (106 molecules per cell ormore) are likely to be monitored if such strains

ever become available.

The NIH program of risk assessment, whichwas formally started in 1979, continues to iden-

tify possible consequences of rDNA research.

Under the aegis of the National Insti tute of Allergy and Infectious Diseases, the program

supports research studies designed to elucidate

the likelihood of harm. * In addition, it collates

general data from other experiments that might

be relevant to risk assessment. Other risk as-

sessments are being conducted by Europeanorganizations * * and by the U.S. EnvironmentalProtection Agency to assess the consequences of

releasing micro-organisms into the environ-

ment.

Thus far, there is no compelling evidence that

E. coli K-12 bacteria carrying rDNA will be more

hazardous than any of the micro-organisms

which served as the source of DNA. Never-

theless, all the experiments have dealt with one

genus o f bacter ium . U nless the conclus ionsabou t E. coli can be extended to other organisms

likely to be u sed in experimen ts (such as Bacillus subtilis and yeast), other assessments maybe ap-propriate.

“Extramural efforts were first conceived in the summer of 1975

to develop and test safer host-vector systems based on E. coli, th einteragency agreement entered into with th e Naval BiosciencesLaboratory tested E. coli systems in a series of simulatedaccidental sp ills in the laboratory. At the University of Michiganthe survival of these systems was tested in m ice and in culturalconditions simulating th e mou se gastrointestinal tract. TuftsUniversity tested these systems in both mice and humanvolunteers.Finally, the survival of host-vector systems in sewagetreatment plants w as tested at the University of Texas. The p eak year for costs of supporting research contracts was 1978; over ahalf-million dollars were required. currently, the cost of maintaining the h igh containment facility at Frederick, Md., isbetw een $200,000 and $250,000 annu ally.

q q First Report to the Com mittee on G enetic Experimentation , ascientific committee of th e International Council of ScientificUnions, from the Working Group on Risk Assessment, July 1978.

Percept ion o f r i sk

The probability of damage can be estimated

for various events. The entire insurance in-

dustry is based on the fact that unfavorable

events occur on a regular basis. The number of

people dying annually from cancer, or automo-

bile accidents, or homicides can be predictedfairly accurately. These est imates depend on

the availability of data and th e assumptions th at

the major determinants do not change fromyear to year.

But even if the probability of damage is fairly

well known, a gap often exists between this

“real” probability of occurrence and the “per-

ceived” probability. Two factors that tend to af-

fect perceptions are the magnitude of the possi-

ble damage and the lack of individual control

over exposure to the risk. Both of these are sig-

nificant factors in the fears associated withrDNA and the manipulation of genes. Because

intuitive evaluations can contradict analyticalevaluations, the question of risk cannot be re-

solved strictly on an analytical basis. Its resolu-

t ion will have to come through the poli t ical

process.

BURDEN OF PROOF

The possibil i ty of inadvertently creating a

dangerous organism does exist, but its prob-

ability is lower than was originally thought.Nevertheless, an important principle emerges

from the debate. Society must decide whetherthe burden of proof rests with those who de-

mand evidence of safety or with those who de-

mand evidence of hazard. The former would

halt experiments until they are proved safe. The

latter would continue experiments until it is

shown that they might cause harm.

A significant theoretical difference exists be-

tween the two approaches. Evidence can almost

a lways be provided to show that somethingcauses harm—e.g., it can be demonstrated that a

poliovirus causes p aralysis, that a Pneumococcuscauses pneumonia, that a rhinovirus causes thecommon cold. However, it cannot be demon-strated that a poliovirus can never cause thecommon cold, It cannot be demonstrated thatrDNA molecules will never be harmful. It can




only be demonstrated that h armful events are level of uncertainty it is willing to accept.

unlikely. Hence, society must determine what

O t h e r c o n c e r n s

C onc e rns ra i se d by indus t r ia l

a p p l i c a t i o n s

Originally concerns involved hazards thatmight arise in the laboratory. Now that thereare ind ustrial ap plications of genetic engineer-ing, the concerns include:

. risks associated with the laboratory con-

struction of new strains of organisms,. risks associated with industrial production

or consumer u se of the new strains, andrisks associated with the products obtained

from the new strains.

Many similar considerations ap ply to the as-sessment of the first two kinds of risks. Unlessthe organisms u sed in an industrial productionscheme are thoroughly characterized, conjec-tured fears about their ability to cause diseasewill continue. Even with a recombinant orga-nism that has a well-defined sequence of DNA, abreak in containment would leave its behaviorin the environment questionable. Experiencewith substances such as asbestos gives rise tofears that exposure to the new biological sys-

tems might also cause unforseen pathologicalconditions at some future time.

Hazards associated with products raise dif-ferent questions. The growing consensus inFederal regulatory agencies appears to be thatthese products should be assessed like allothers—e,g., human growth hormone (hGH)produced by genetically engineered bacteriashould be tested for purity, chemical identity,and biological activity just like hGH from h um anpituitary glands. The possibility of productvariation due to mutation of the bacteria,

however, suggests that batch testing and certifi-cation m ight be war ranted as well. (For furtherdiscussion see ch. 11.)

C onc e rns ra i se d by the imp l i c a t ions o f

the rDNA controversy for general

m i c r o b i o l o g y

Questions about the potential harm fromgenetically engineered micro-organisms haveled to questions about the efforts currentlyemp loyed to protect the pu blic from work beingdone w ith micro-organisms known to be hazard-ous. These viruses, bacteria, and fungi arehand led daily in laboratory experiments, in theroutine isolation of infectious agents from pa-tients, and in the pr odu ction of vaccines in thepharmaceutical industry.

Questions have been raised about the efficacyof regulations established for these variouspotentially hazardous agents. A full-scale assess-ment is not within the scope of this study, but itis clear that the questions are pertinent. Twoconclusions have been reached.

First, there is a grow ing belief that th e mereexistence of a classification scheme for hazard-ous agents by the Center for Disease Control

(CDC) is not enough to ensure their safe han-

dling. The Subcommittee on Arbovirus Labora-tory Safety was formed recently because of con-cerns expressed in academic circles. Represent-atives from universities, the Public Health Serv-ice, the U. S. Department of Agriculture, andthe military, who constituted the subcommit-tee, are preparing a report based on an interna-tional survey of laboratory p ractices and infec-tions. They found wide variation in the waysdifferent agents were handled. Most of theirrecommendations are identical with those ap-plicable to rDNA–that appropriate containmentlevels be used with different viruses, that the

health of workers be monitored, and that an In-stitutional Biosafety Committee be appointed toserve each institution.




Second, little is known about the healthrecord of workers involved in the fermentationand vaccine industries. For most industrialoperations the evidence of harm is almost en-tirely anecdotal. Most ind ustrial fermentationsare regarded as harmless; representatives of in-dustry characterize it as a “non-problem” thathas never merited monitoring. Comprehensiveinformation on the potential harmful effectsassociated with research using rDNA-carrying

. micro-organisms will not be available becausethe Gu idelines consider it the respon sibility of each institution or company to “determine, inconnection with each project, the necessity formedical surveillance of recombinant-DNA re-search personnel. ” Hence some institutionsmight d ecide to keep r ecords of some or all ac-tivities; others m ight not.

To be sure, some companies have exceeded

the minimal medical standards set by NIH forfermentation using rDNA-carrying micro-orga-nisms–e.g., Eli Lilly & Co. requires that allillnesses be reported to su pervisors and that anyemployees who are ill for more than 5 daysmu st report to a physician before being allowedto return to w ork. Any employee taking antibi-otics (which might make it easier for bacteria tocolonize) is restricted from areas w here rDN Aresearch is being done until 5 days after the dis-continuance of the antibiotic. At Abbott Labora-tories, a physician checks into the illness of anyrecombinant worker who is off more than 1day–a precaution taken only after 5 days off for workers in other areas. Lilly maintains acomputer listing of all workers involved inrDNA activities. Lilly, the Upjohn Co., andMerck, Sharp and Dohme have been in theprocess of computerizing the health records of all their em ployees over the p ast several years.

Work with rDNA has focused attention onbiohazards an d m edical surveillance—an aw are-ness that had a risen in the past but h ad n ot beensustained. * Consequently, several documentson the subject either ha ve been or w ill be pu b-

lished:

q As of September 1980, the National Institutes of occupationalSafety and Health and the Ent’ironnlental Protection Agency were

planning to fund assessments of the adequacy of current medical

surveillance technology,

q

q

q

q

CDC is preparing a complete revision of itslaboratory safety manual, which is widelyused as a starting point by other labora-tories.The Classification of Etiologic Agents on the

Basis of Hazard, which was last revised in1974, has been expanded by CDC in collab-oration with NIH into a Proposed BiosafetyGuidelines for Microbiological and Biomedi-

cal Laboratories. These guidelines serve thepu rpose fulfilled by the Dangerou s Patho-gens Ad visory Group (DPAG) in the UnitedKingdom, although they lack any regula-tory strength.A comprehensive program in safe ty,health, and environmental protection wasdeveloped in 1979 by and for NIH. It is ad-ministered by the Division of Safety, whichincludes programs in radiation safety, oc-

cupational safety and h ealth, environm en-tal protection, and occup ational med icine.The Office of Biohazard Safety, NationalCancer Insti tute has just completed a 3-

year study of the medical surveillance pro-

grams of its contractors; a report is being

drafted.

Although the academic, governmental, andindustrial communities have shown growing in-terest in biosafety, * no Federal agency regulatesth e possession or use of micro-organisms except

for those highly pathogenic to animals and forinterstate transport. * * Whether such regula-

tions are necessary is an issue that extends be-yond the scope of this study. Nevertheless,

other countries—for instance the United King-

dom, with its DPAG—have acted on the issue.

This organization functions specifically to guard

against hazardous micro-organisms, by moni-

toring and licensing university and industriallaboratories and meting out penalt ies when

necessary.

“Curiously, there is no formal society or journal, hut there has

been an annual Biologica l Safety Conference s ince 1955, con-

ducted on a round-rohin hasis primarily h~ close associates of thelate Arnold G. Wedum, M. D.—fornler Director of Industrial Health

anci Safet-v at the LI. S. Army Bioiogica] Research Laboratories, FortDetrick , Md., who is regarded as the “Father of MicrobiologicalSafety. ”

q “In some States and cities, licensing is required for all facilitieshandling pathogenic micro-organisms.




C onc e rns ra i se d by the imp l i c a t ions o f outside the cell. If the DNA is combined within

the rDNA controversy for o ther living cells, the Guidelines d o not per tain. Figur e

g en e t i c m a n i p u l a t i o n 35 shows several methods that achieve the samegoal–transfering genetic material from on e cell

Alter ing the hereditary character is tics of an to another, bypassing the normal sexualorganism-by using

several methods of definition of rDNAcombination of the

rDNA is just one of the mechanisms of ma ting. It is par ticularly signifi-

genet ic engineer ing. The cant that DNA from different species can berefers specifically to the combined by all these mechanisms, only one of DNA from two organisms which is rDNA. Different species of bacteria,

Figure 37.-Alternative Methods for Transferring DNA From One Cell to Another

A. The two cells are fused in totoB. A microcell with a fragmented nucleus carries the DNAC. Free DNA can enter the recipient cell in a number of ways: by direct microinjection, through

calcium-mediated transformation, or by being coated with a phospholipid membrane inorder to fuse with the recipient cell

D. The free DNA can be joined to a plasmid and transferred as recombinant DNA




Ch. 10—The Question of Risk q 207

fungi, and h igher organisms can all be fused ormanipulated. *

Opponents of rDNA have stated that combin-ing genes from different species may disturb anextremely intricate ecological interaction that isonly dimly understood. Hence, such experi-ments, it is argued, are unpredictable and there-fore hazardous. If so, all the other methodsrepresented in figure 35 should be included inthe Guidelines. Yet they are not.

The most acceptable explanation for this in-consistency is that rDNA is currently the most

q For example, antibiotic resistant plasmids have been trans-ferred from Staphylococcus aureus to Bacillus subtilis acrossspecies barriers by transformation, not by rD NA. Foreign genesfor the enzyme amylase have also been introduced into B. subtilis.

Conc lus ion

Thus far, no demonstrable harm associatedwith genetic engineering, and particularlyrDNA, has been found , But although d emonstra-ble harm is based on evidence that dam age hasoccurred at one t ime or another, i t does not

mean that damage cannot occur.

Conjectural hazards based on analogies and

scenarios have been addressed and most have

proved l e s s w or r i som e than p rev ious ly a s -

sumed. Nevertheless, there is agreement that

certain experiments, such as the transfer of genes for known toxins or venoms into bacteria,

should still be prohibited because of the real

l ikelihood of danger. Sti l l other experiments

cannot clearly be shown to be hazardous or

readily dismissed as harmless. Hence, a political

decision is likely to be required to establish

w ha t cons t i t u t e s accep tab le p roof and w ho

must p rovide it.

Given that potential harm can be identified in

some cases, its probable occurrence and magni-

tude quantified, and perceived risk taken into

account, a decision to proceed is usually basedon society’s willingness to take the risk. This

triad of the physical (actual risk), psychological

(perception of risk), and political (willingness to take risk) plays a role in all decisions relating togenetic engineering.

efficient and successful method of combininggenes from very diverse organisms. It is reason-able to ask, however, what would happen if anyof the other methods become equally success-ful. Will a profusion of guidelines appear? Willone committee oversee all genetic experiments

E th ic a l an d m ora l conc erns

The perceived risk associated with geneticengineering includes ethical and moral hazardsas well as physical ones. It is important torecognize that these are part of the generaltopic of risk. To some, there is just as much risk to social values and structure as to humanhealth and the environment. (For further dis-cussion see ch. 13.)

The potential benefits must always be con-sidered along w ith the risks. Decisions m ade byRAC have reflected this view—e.g., when itapp roved th e cloning of the genetic material of the foot-and -mouth disease virus. The perceivedbenefits to millions of animals outweighed thepotential hazard.

Recombinant DN A techniques repr esent justone of several methods to join fragments of

DNA from different organisms. The currentGuidelines do no extend to these other tech-niques, although they share some of the sameun certainties. Ignoring the consequences of theother technologies might be viewed a s an incon-sistency in policy.

While the initial concerns about the possibili-ty of hazards at the laboratory level appear tohave been overstated, other types of potentialhazards at different stages of the technologyhave been identified. Emphasis has shiftedsomewhat from conjectured hazards that might

arise from research and development to thosethat might be associated with production tech-nologies. As a consequence, there is a clearermandate for existing Federal regulatory agen-cies to play a role in ensuring safety in industrialsettings.



C h a p t e r 11

R e g u la t i on o f

G e n e t i c E n g i n e e r i n g



C h a p t e r 11

Page

[introduction . . , . . . . . , . . . . , . . . . , . . . . . . . . , 211Framew ork for t he Analys is. . . . . . . . . . . . . . .211

Curren t Regulation: the NIH Gu idelin es. . . . . . . , 212

Substantive Requirements . . . . . . . . . . . . . . . .212

Administration . . . . . . . . , . . . . . . . . . . . . . .212Provisions for Voluntary Compliance . . , . . .215Evaluation of the Guidelines . , . . . , . . . . . , . . . 216

T h e P r o b l e m o f Risk . . . . . . . . . . . . . . . . . . .216The Decision making Process . . . . . . . . . . . . .221

Conclusion. . . . . . . . . . . . . . . . . . . . . . . . . . . .223oth er M ean s of Regu latio n . . . . . . . . . . . . . . . . ..224

Federal Statutes. . . . . . . . . . . . . . . . . . . . . . . .224“T’ort Law and Workm en’s Compen sation . . , . .227

State and Local Law . . . . . . . . . . . . . . . . . . . . , 229

Conclusion . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 230Issue and options . . . . . . . . . . . . . . . . . . . . . . . . 230

Tables

Table No. P a g e

35. Containment Recommended by NationalInstitutes of Health . . . . . . . . . . . . . . . . . . . .,213

36. Statutes That Will Be Mo st App licable? to

Commercial Genetic Engineering. . . . . . . . . . 224



Cha p t e r 11

R e g u l a t i o n o f G e n e t i c E n g i n e e r i n g


Although no evidence exists that any harmful

organism has been created by molecular genetictechniques, most experts believe that somerisk* is associated with genetic engineering.One kind is relatively certain and quan tifiable—that of working with known toxins or patho-gens. Another is uncertain and hypothetical–that of the p ossible creation of a pathogenic orotherwise undesirable organism by reshufflinggenes thought to be harmless. These may bethought of as physical risks because they con-cern hu man h ealth or the environment.

Concern has also arisen about the possiblelong-range imp acts of the techniqu es—that th eymay eventually be used on humans in somemorally unacceptable manner or may changefundam ental views of what it means to be hu -man. These possibilities may be thought of ascultural risks, since they threaten fundamentalbeliefs and value systems.

1

The issue of whether or not to regulatemolecular genetic techniques—and if so, towh at extent—defies a simp le solution. Percep-tions of the nature, magnitude, and acceptabili-ty of the risks differ d rastically. Appr oximately6 years ago, when the scientific community it-

self accepted a moratorium on certain classes of recombinant DNA (rDNA) research, some sci-

entists considered the concern unnecessary. To-

day, even though the physical risks of rDNA re-search are generally considered to be less thanoriginally feared—and the realization of i ts

benefits much closer–some people would stillprohibit it.

The Federal Government’s approach to this

issue has been the promulgation of the Guide-lines for Research Involving Recombinan t DN AMolecules (Guidelines), by the National Insti-

tutes of Health (NIH). (See app. III-C for infor-mation about what other countries have done

q As used in this chapter, risk means th e possibility of harm. ‘I-heprobability ot that harm occurring may he extremely low anckr

highly uncertain.‘H. “Ilistam, f tngelhard t , Jr. , ‘Takin g Risks: Some Background

Issues in th e I)etxite (k)ncern ing Recombinant DNA Research,Southern (;al(~ornia Law Review .51:6, pp. 1141-1151, 1978.

with respect to guidelines for rDNA.) Three

other available modes of oversight or regulationare curren t Federal statutes, tort law , and Stateand local law.

F r a mew o r k f o r t h e a n a l y s i s

In deciding how to address the risks posed bygenetic engineering, some of the importantquestions that need to be examined are:

. How broadly the scope of the issue (orproblem) should be defined.—Who identifies the risks and their mag-

nitude?—Who proposes the means for addressing

the p roblem?q The nature of the procedural, decisionmak-

ing mechanism.—Who decides?—Who will benefit from the p roposed ac-

tion and w ho w ill bear the risk?—Will the risk be borne voluntarily or in-

voluntarily?—Who has the burd en of proof?—Shou ld a risk/ benefit analysis, or some

other approach, be used?q The available solutions and their adequacy.

—Should there be full regulation, no reg-ulation, or something in-between?

—What actions and actors should be cov-‘ered?

—What is the appropriate means for en-forcing a regulatory decision?

–Which agency or other group should dothe regulating?

Und erlying these questions is the prop osition,widely accepted by commentators on sciencepolicy, that scientists are qualified to assessph ysical risk, since that involves measur ing andevaluating technical data. However, a judgment

of safety (the acceptability of that risk) can onlybe made by society through the political proc-ess, since it involves weighing and choosingamong values.

z 3456 Scientists are not nec-

‘William W. I ,owrance , Of Aceep(able Risk: tkience and the De-

termination of &fety (l,os Altos, (hl i t . : wi]l ian l Kau fmann , ]n(~.,

1976).

211



212 . impacts of Applied Genetics—Micro-Organisms, Plants, and An/reals

essarily considered to be more qualified to make high value they place on unrestricted research

decisions concerning social values than other and because of possible conflicts of interest.

well-informed persons; they m ay in fact be less Moreover, according to this view, if society is to

qualified when the decision involves possible bear a risk, it shou ld jud ge the acceptability of

restrictions on scientific research because of the that risk and give its informed consent to it.7*

((:f)ntilll:~!fl,f}olll p. 211)‘tAlvin W. Wcinlwrg , “Scienct~ i l l l [l ‘1’l’illlS-SCit? ll(X?, ” A4inerva,

10:2, April 1972. 7Engelhard t , op.cit .; Lowrance , op.cit .; an d Bazelon, op. cit.

4Allan Mazur, ‘(Disputes Between Kxperts, ” Minerva 11:2, April q [n practice, it may often he difficult to keep the two kinds of1973. decisions separate, since the values of individual scientists may in -

‘A1.thur Kantrowitz, ‘t’r’he Scien ce Court Experim ent, ” Juri- tluence their interpretation of technical data, and since policy-

metrics ./ourna/, vol. 17, 1977, p. 332. makers may not have th e technical competence to understand the‘David L.Bazelon, “Risk and Responsibility, ” Science, vol. 205, risks sufficiently.e

July 20, 1979, pp. 277-280. ‘Weinb erg, op. cit.; and Bazelon, op. cit.

C u r r e n t r e g u l a t i o n : t h e N I H G u i d e l i n e s

The Guidelines have been developing in

stages over a period of approximately 6 years asscientists and policymakers have grappled withthe risks posed by rDN A techniques. (This his-tory, discussed in app. III-A, is crucial to under-standing current regulatory issues, and it servesas a basis for evaluating the Gu idelines.) Theyrepresent the only Federal oversight mecha-nism that specifically addresses genetic engi-neering.

Subs tan t i v e re qu i re me n t s

The Guidelines apply to all research involvingrDNA molecules in the United States or its ter-ritories conducted at or sponsored by any in-stitution receiving any support for rDNA re-search from NIH. Six types of experiments arespecifically prohibited: 1) the formation of rDNA derived from certain pathogenic orga-nisms; 2) the formation of rDNA containinggenes that make vertebrate toxins; 3) the u se of the rDNA techniques to create certain plantpathogens; 4) transference of drug resistancetraits to micro-organisms that cause disease inhumans, animals, or plants; 5) the deliberate

release of any organism containing rDNA intothe environment; and 6) experiments usingmore than 10 liters (1) of culture unless therDNA is “rigorously characterized and theabsence of harmful sequences established. ” Aprocedu re is specified for obtaining exceptions

from these prohibitions. Five types of experi-

ments are completely exempt.

Those experiments that are neither prohib-ited nor exempt must be carried on in ac-cordance with physical and biological contain-ment levels that relate to the d egree of potentialhazard. (See table 35.) Physical containment re-quires methods and equipment that lessen thechances that a recombinant organ ism might es-cape. Four levels, designated P1 for the leastrestrictive through P4 for the most, are defined.

Biological containment requires working withweakened organisms that are unlikely to sur-

vive any escape from the laboratory. Threelevels are specified. Classes of permitted ex-periments are assigned both physical and bio-logical containment levels. Most experimentsusing Escherichia coli K-12, the standard labora-tory bacterium used in approximately 80 per-cent of all experiments covered by the Guide-lines, may be p erformed at the lowest contain-ment levels.

ADMINISTRATION

The Guidelines provide an administrativeframework for implementation that specifies

the roles and responsibilities of the scientists,their institutions, and the Federal Government.The parties who are crucial to the effectiveoperation of the system are: 1) the Director of NIH, 2) the NIH Recombinant DNA AdvisoryCommittee (RAC), 3) the NIH Office of Recombi-



Ch. 11—Regulation of Genetic Engineering q 213

Table 35.–Containment Recommended byNational Institutes of Health

Biological—Any combination of vector and host must bechosen to minimize both the survival of the systemoutside of the laboratory and the transmission of thevector to nonlaboratory hosts. There are three levelsof biological containment:

H V 1– Requires the use of Escherichia coli K12 orother weakened strains of micro-organisms thatare less able to live outside the laboratory.

HV 2– Requires the use of specially engineered strainsthat are especially sensitive to ultraviolet light,detergents, and the absence of certainuncommon chemical compounds.

H V 3– No organism has yet been developed that canqualify as HV3.

Physical—Special laboratories (P1-P4)

P1— Good laboratory procedures, trained personnel,wastes decontaminated.

P2— Biohazards sign, no public access, autoclave inbuilding, hand-washing facility.

P3 – Negative pressure, filters in vacuum line, class IIsafety cabinets.

P 4 – Monolithic construction, air locks, all airdecontaminated, autoclave in room, allexperiments in class Ill safety cabinets (glovebox), shower room.


nant DNA Activities (ORDA), 4) the Federal In-

teragency Advisory Committee on Recombinant

DNA Research (Interagency Committee), 5) theInstitution where the research is conducted, 6)the Institutional Biosafety Committee (IBC), 7)

the Principal Investigator (PI), and 8) the Bio-

logical Safety Officer.

Th e Director of NIH carries the primary bur-den for the Federal Governm ent’s oversight of rDNA activities, since he is responsible for im-plementing and interpreting the Guidelines, es-tablishing and maintaining RAC (a technical ad-visory committee) and ORDA (whose functionsare purely administrative), and maintaining theInteragency Committee (which coordinates allFederal activities relating to rDN A). Under th isarrangement, all decisions an d actions are taken

by the Director or his staff. For major actions,the Director must seek the ad vice of RAC, andhe mu st provide the pu blic and other Federalagencies with at least 30 days to comment on

proposed actions. Such actions include: 1)assigning and changing containment levels forexperiments, 2) certifying new host-vector sys-tems, 3) maintaining a list of rDNA molecules ex-empt from the Guidelines, 4) permitting excep-tions to prohibited experiments, and 5) adop ting

changes in the Guidelines.

For other sp ecified actions, the Director needonly inform RAC, the IBCs, and the p ublic of hisdecision. The most important of these are: 1)making m inor interpretive decisions on contain-ment for certain experiments; 2) authorizing,under procedures specified by RAC, large-scalework (involving m ore than 101 of culture) withrDN A that is rigorously characterized and freeof harmful sequences; and 3) sup porting labora-tory safety training programs. Every actiontaken by the Director pursuant to the Guide-

lines must present ‘(no significant risk to healthor the environment. ”

RAC is an advisory committee to the Directoron technical matters. It meets quarterly. Its pur-pose, as described in its current charter of Jun e26, 1980 (and unchanged since its inception inOctober 1974), is as follows:

The goal of the Comm ittee is to investigatethe current state of knowledge and technologyregarding DN A recombinant, their survival innature, and transferability to other organisms;to recomm end guidelines for the condu ct of

recombinant DN A experiments; and to recom-mend programs to assess the possibility of spread of specific DNA r ecombinant and thepossible hazards to public health and to the en-vironment. This Committee is a technical commit-

tee, established to look at a specfic problem. (Em-phasis added.)

The charter and the Guidelines also assign it

certain advisory functions that have changedover time.

The RAC is composed of not more than 25members. At least eight must specialize in mo-

lecular biology or related fields; at least six must

be authorities from other scientific disciplines;and at least six must be authorit ies on law,

public policy, the environment, public or oc-

cupational health, or related fields. In addition,

76-565 0 - 81 - 15




representatives from various Federal agencies

serve as nonvoting members.

ORDA performs administrat ive functions,

which include reviewing and approving IBC

membership and serving as a national center

for information and advice on the Guidelinesand rDNA activities.

The Interagency Committee was established inOctober 1976 to advise the Secretary of the thenDepartment of Health Education and Welfare(HEW) [now Health and Human Services(DHHS)] and the Director of NIH on the coor-dination of all Federal activities relating torDN A. It has thu s far prod uced tw o reports. Itsfirst, in March 1977, concluded that existingFederal law w ould n ot permit the regulation of all rDNA research in the United States to the ex-tent considered necessary and recommendednew legislation, specifying th e elements of thatlegislation. ’” The second, in November 1977,surveyed international activities on regulatingthe research and concluded that, while appro-priate Federal agencies should continu e to w ork closely with the various international organiza-tions, no formal governm ental action w as neces-sary to prod uce internationa l control by meansof a treaty or convention.

11It is currently con-

sidering issues arising from the large-scale in-du strial applications of rDNA techniques.

Under the Guidelines, essentially all the re-

sponsibility for overseeing rDNA “experimentslies with those sponsoring or condu cting th e re-search. The Institution must implement generalsafety policies, * establish an IBC, which meetsspecified requirements, and appoint a BiologicalSafety Officer. The Biological Safety Officer, whois needed only if the Institution conducts ex-periment s requiring P3 or P4 containment , (seetable 35) oversees safety standards. The initialresponsibility for particular experiments lies

‘Interim Report of the Federal Interagency Committee on Recom- binant DNA Research: Su,gested Elements-for Legislation, Mar. 15,

1977, pp . 9-10.ID lb id . , pp . 11-15,I I Report of th e Federa l intet-~ency Committee on Recombinant

DNA ftesearch: International Activities, Novendxx 1977, pp. 13-15.“These include conducting im y heid[h surveillance that it deter-

mines to be necessary an d ensuring iippr~priiiie tritining for th eIBC , Biological Safety officers, Principal lnv{?sti~iit~rs, an d labora-tory staff.

with the PI, the scientist receiving th e fund ing.This person is responsible for determining andimplementing containment and other safe-guard s and training and supervising staff. In ad-dition, the PI must also submit a registrationdocument that contains information about thepr oject to the IBC, and pet ition N IH for: 1) cer-tification of host-vector systems, 2) exceptionsor exemptions from the Guidelines, 3) and de-termination of containment levels for experi-ments not covered by the Guidelines. Further-more, all of the above have certain r eporting re-quirements d esigned so that ORDA is eventuallyinformed of significant problems, accidents, vio-lations, or illnesses.**

The IBC is designed to p rovide a quasi-ind e-pendent review of rDNA work done at an in-stitution. It is responsible for: 1) reviewing all

rDNA research conducted at or sponsored bythe institution and approving those projects inconformity with the Guidelines; 2) periodicallyreviewing ongoing p rojects; 3) adop ting emer-gency plans for spills and contamination; 4)lowering containment levels for certain rDNAand recombinant organisms in which the ab-sence of harmful sequences has been estab-lished; and 5) reporting significant problems,violations, illnesses, or accidents to ORDAwithin 30 days.*** The IBC mu st be comprisedof no fewer than five members who can col-lectively assess the risks to health or the en-

vironment from the experiments. At least 20percent of the membership m ust not be other-wise affiliated with the institution where thework is being done, and mu st represent the in-terests of the surrounding community in pro-tecting health and the environment. Comm-ittee members cannot review a project inwh ich th ey have been, or expect to be, involvedor have a direct financial interest. Finally, theGuidelines suggest that IBC meetings be public;minutes of the meetings and submitted docu-ments must be available to the public onrequest.

q q The PI is required to report this information with in 30 days toORDA an d his IBC . The Biological Siifet~ officer mus l report thesame to the Institution and the IBC unless the PI h as clone so. Theinstitution must report within 30 d i i ~ s t o ORDA unless the PI orIB(: ha s done so.

*” “It does no t have to report if the PI h i t s done SO .




Ev a lu a t ion o f t he Gu ide l ine s

Two basic issues mu st be ad dressed. The firstis how well the Guidelines confront the risksfrom genetic engineering, wh ich m ay not h ave adefinitive answer in view of the uncertainty

associated with most of the risks. Consequently,it is also necessary to consider a second issue—whether confidence is warranted in the deci-sionmaking p rocess responsible for the Gu ide-lines.

THE PROBLEM OF RISK

The Guidelines are designed to address therisks to public health and the environment fromeither rDNA molecules or organisms and vi-ruses containing them. The underlying premiseis that research should not be unreasonablyrestricted. This is essentially a risk-benefit ap-

proach; at the time that the original Guidelineswere drafted, it represented a compromise be-tween the extremes of no regulation and of noresearch without proof of safety. physical andbiological containment levels were establishedfor various experiments based on estimateddegrees of risk. The ad ministrative mechanismcreated by the Guidelines is that of a Federalagency —NIH—advised by a diverse body of experts— RAC. Scientific advice on the technicalaspects of risk assessment is provided by techni-cal experts on RAC; public input is provided byexperts in nontechnical subjects and by the

right of the public to comment on major actions,which are published in the Federal Register.Compliance is accomplished by a combinationof local self-regulation and limited Federal over-sight, with the ultimate enforcement resting inthe Federal funding p ower.

Since their initial app earance, the Gu idelineshave evolved. As scientists learned more abou trDNA and molecular genetics, two trends oc-curred. First, containm ent levels were p rogres-sively lowered. Major revisions were made in1978 and 1980; minor revisions were often

made quarterly, as proposals were submitted tothe RAC at its quarterly meetings, recom-mended by RAC, and accepted by the Director.By now, approximately 85 percent of the per-mitted experiments can be done at the lowestphysical and biological containment levels. Se-cond, the degree of centralized Federal over-

sight has been substantially red uced to the pointwhere almost none remains. Under the 1976Guidelines, all perm itted experiments u ltimatelyhad to be reviewed by the IBC and ORDA beforethey could be started; the 1978 Guidelines nolonger required preinitiation review of most

experiments by ORDA, although ORDA con-tinued to maintain a registry of experimentsand to review IBC decisions. Under theNovember 1980 revision to the Guidelines, therewill be no Federal registration or review of ex-periments for which containment levels arespecified in the Guidelines. About 97 percent of the permitted experiments fall into thiscategory.

Preinitiation review of experimen ts by RAChas been an important part of the oversightmechanism. Expert review encourages experi-

mental design to be well thought out an d pro-vides a m eans for catching potential pr oblems,e.g., one application reviewed by RAC nevermentioned th at the species to be used as a DN Adonor was capable of manu facturing a potentneurotoxin; it was turned down after a RACmember familiar with the species brought thisfact to the Committee’s attention.

13

The burdens imposed on rDNA activities bythe Guid elines app ear to be reasonable in viewof continu ing concerns abou t risk. Less than 15percent of permitted experiments require pre-

initiation app roval by th e local IBC’s, wh ich usu -ally meet monthly. Preinitiation approval of ex-periments by NIH is required only for: 1) experi-ments that have not been assigned containmentlevels by the Guidelines; 2) experiments usingnew host-vector systems, wh ich m ust be certi-fied by NIH; 3) certain experiments requiringcase-by-case approval; and 4) requests for ex-ceptions from Guideline requirements. The low-est containment levels place minimal burdenson the experimenter. (see table 35). For in-dustrial applications, NIH approval must bereceived n ot only w hen th e project is scaled-up

beyond the 10-1 limit, but also for each additional scale-up of the same p roject. Many representatives of industry consider these subse

‘3R. M. Henig, “Trouble on th e RAC—Committee Splits OveDowngrading of E. co/i Containment,” Bioscience, vol. 29, p p. 7 5 9

762, December 1979.



Ch. n-Regulation of Genetic Engineering q 217

quent ap provals to be unnecessary and burden-some.

Information about whether the Guidelineshave been a disadvantage for U.S. companies ininternational competition is scanty. Examplesinclud e the approximately l-year headstart tw o

European grou ps w ere given wh ile the cloningof hepatitis B virus was prohibited, the advan-tage some European companies had in usingcertain species of bacteria for cloning underconditions that were prohibited in the UnitedStates, and the delays some pharmaceuticalcompanies faced because they had to build bet-ter containment facilities.

The present Guidelines are a comprehensive,flexible, and nonburdensome way of dealingwith the physical risks associated w ith rDNA re-search while permitting the work to go for-

ward . That is all they were ever intend ed to do.

The Scope of the Guidelines.—In manyrespects, the Gu idelines do not address the fullscope of the risks of genetic engineering. Theycover one technique, albeit the most important;they do n ot add ress the admittedly uncertain,long-term cultural risks; they are not legally

binding on researchers receiving funds fromagencies other than NIH; and they are not bind-ing on industry.

Other genetic techniques present risks simi-lar to those posed by rDNA, but to a lesser de-

gree. Recombinant DNA is the most versatileand efficient technique; it uses the greatestvariety of genetic material from the widestnumber of sources with reasonable assuranceof expression by the host cell. Cell fusion of micro-organisms, which also involves the un cer-tain risk of recombining the genetic material of different species, is significantly less versatileand efficient than rDN A but mixes more geneticmaterial. In add ition, the pa rental cells may con-tain partial viral genomes that could combine toform a complete genome when the cells arefused. Transformation, a technique known fordecades, similarly involves moving pieces of DNA between different cells. However, it is sig-nificantly less versatile and efficient th an cell fu-sion, and it is generally considered to be virtual-ly risk-free. Thus, cell fusion is in a gray area

between the other two techniques; yet no risk assessment h as been don e, and no Federal over-sight exists.

Another limitation in the scope of the guide-lines—and in the process by which they were

formulated—is that long-range cultural risks (as

distinguished from policy issues related to safe-ty) were never addressed. As noted by the Di-rector of NIH:

14

NIH has been addressing the policy ques-tions involving the safety of this research, notthe ‘potential future ap plication . . . to the alter-ing of the genetic character of higher forms of life, including man’ . . .

Perhaps it was inappropriate to do more. Suchethical issues might be considered premature inview of the level of the development of the tech-nology. The desire among many molecular bi-

ologists to move ahead with the research meantthat experiments were being done; thereforethe immed iate potential for harm was to healthand the environment. Thus, it was arguablynecessary to develop a framework to deal withthe risks based on what was known at the time.On the other hand, the broader questions of where the research might eventually lead andwhether it should be done at all have beenraised in the p ublic debate. They hav e not beenformally considered by the Federal Govern-ment.

Another limitation in the scope of the Guide-lines is their nonapplicability to researchfunded or performed by other Federal agencies.However, agencies supporting such researchare complying with the Gu idelines as a m atter of policy. There appears to be little reason forquestioning these declarations of general policy.In practice, problems might arise if a mission isperceived to be at odds with the Guidelines orbecause of simple bureaucratic defense of terri-tory —e.g., when the 1976 Guidelines were pro-mu lgated, two agencies—the Dep artment of De-

fense (DOD) and the N ational Science Founda-

tion (NSF) preserved the right to deviate for rea-sons of national security or differing interpreta-

1 4 4 3 F.R. 6o103 , Dec. 22, 1978, c i t ing 43 F.R. 33067 , JuI.v 28 ,

1978.



220 q Impacts of Applied Genetlcs—Micro-Organisms, Plants, and Animals

sight. Penalties are based on N IH’s pow er to re-strict or terminate its funding.

The initial responsibility for compliance lieswith the scientist doing the experiments. A re-searcher’s attitude toward the risks of rDNAtechniqu es and the necessity for the Guid elines

app ear to be an influential factor in the degreeof compliance. A science writer who worked for3 months in a un iversity lab in 1976 noted slop-

py procedures and a cavalier attitude, stat ing:“Among the young graduate students and post-

doctorates it seemed almost chic not to know

the NIH rules.”22

On the other hand, in the caseof a recent violation of the Guidelines, it appears

as if the investigator’s graduate students were

the first to raise questions.23 24 C om pet i t i veness

is another important factor. Novice scientists

must establish reputations, secure tenure in a

t ight job market, and obtain scarce research

funds; established researchers still compete forgrants and certainly for peer recognition. This

competitive pressure could provide strong in-

centives to bend the Guidelines; on the otherhand, it might be channeled to encourage com-

pl iance i f it i s bel ieved that NIH wi ll in fact

penalize violations by restricting or terminating

funding.

The first level of actual oversight occurs atthe institution. An argument can be mad e thatreliance on the PI and an IBC (that might becomposed mostly of the PI’s colleagues) provides

too great an opportunity for lax enforcement orcoverups. On the other hand, spreading respon-sibility among the institution, the PI, the IBC,and, in the case of more hazardous experi-ments, the Biological Safety Officer might re-du ce the chance of violations being overlookedor condon ed. This responsibility is enhanced bythe reporting requirements borne by each of these parties, designed so that O RDA learns of “significant” problems, acciden ts, violations, andillnesses. What is “significant” is not defined .

Pub lic involvement at th e local level acts as anadditional safeguard. Twenty percent of the

~ZJane t L. Hopson, “Recombinan t Lab for DNA an d MY 95 Days

in It,” Smithsonian, vol. 8, June 1977, p. 62.Z3D0 Di cks on , (, Another Vio]ation of N IH Guide] ines, j) ~a t u r e vol.

286, Aug. 14, 1980, p. 649.i~D. Di cks on , “DNA Recomb in a t i o n Forces Resignation , ” ~tlfUJl?

vol. 287, Sept . 18 , 1980, p . 179.

IBCs members m ust be u naffiliated with the in-stitution. IBC documents, includ ing m inutes of meetings, are publicly available, but meetingsare not required to be held in public. On theother hand, the probable inability of the mem-bers who represent the public to understandthe technical matters might limit their effective-

ness.

How successful h as compliance been? Threeknown violations have occurred. In each, nothreat to health and the environment existed. Ineach, there was some confusion as to why theviolations occurred. NIH is presently invest-igating the third violation. For the first two, itaccepted explanations of misunderstandingsand misinterpretations of the Guidelines. How-ever, a Senate oversight report concluded:

25

While und oubtedly most researchers haveobserved the gu idelines conscientiously, it is

equally clear that others have su bstituted theirown judgments of safety for those of NIH.

No firm conclusions can be draw n on the ques-tion of compliance. The rep orting of on ly a fewviolations could be evidence that th e comp liancemechanism embodied in the Guidelines hasbeen working w ell. Or it could mean th at someviolations are not being discovered or rep orted.

The November 1980 amendments to theGuidelines substantially changed proceduresdesigned to monitor compliance by abolishing adocument called a Memorandum of Under-

standing an d Agreem ent (MUA). It had been re-quired for 15 to 20 percent of all experiments,those thou ght to be p otentially most risky. TheMUA, which w as to be filed w ith ORDA by aninstitution, provided information abou t each ex-perimen t, and it was the institution’s certifica-tion to NIH that the experiment complied withthe Guidelines. By having the MUAs, ORDAcould monitor for inconsistencies in interpret-ing the Guidelines, actual noncompliance, andthe consistency and quality with which IBCsfunctioned nationwide. The amendments con-tinued a trend begun in January 1980, when ap-

proximately 80 percent of the experiments,

‘s’’ Recombinant D N A Research an d Its Applications,” Oversight

Report, Subcommittee on Science, Technology, and Space of theSenate Committee on Commerce, Science and Transportation,Aug . 1978, p. 17.



Ch. 11-Regulation of Genetic Engineering . 221

those done with E. coli K-12, were exemptedfrom the MUA requirement.

The abolition of the MUA essentially abol-ished centralized Federal monitoring of rDNAexperiments. The only current Gu ideline provi-sion that serves this kind of monitoring function

is the requ irement that the institu tion, the IBC,or the PI notify ORDA of any significant viola-tions, accidents, or problems with interpreta-tion. Limited monitoring of large-scale activitiescontinues. Under NIH procedures (which arenot p art of the Guidelines) for reviewing ap pli-cations for exemptions from the 10-1 limit, theapp lication m ust include a copy of the registra-tion document filed with the IBC. The manufac-turing facilities may also be inspected by N IH,not for regulatory purposes, but to gather infor-mation for updating its recommended large-scale containment levels; The abolition of the

MUA is consistent with traditional views thatGovernment should not interfere with basic sci-entific research. Whether or not it will reduceeither the incentive to comply with the Guide-lines or the likelihood of discovering violationsremains to be seen.

THE DECISIONMAKING PROCESS

Another way to evaluate the Guidelines be-sides considering their substantive require-ments is to look at the process by which theywere formulated. In a situation where there isuncertainty and even strong disagreement

about the n ature, scope, and m agnitude of therisks, it is difficult to judge whether or not aprop osed solution to a problem will be a goodone. Society’s confidence in the decisionmakingprocess and in the decisionmakers then be-comes the issue. As David L. Bazelon, Chief Judge of the U. S. Court of Appea ls for the Dis-trict of Columbia, has stated:

26

When th e issues are controversial, any deci-sion may fail to satisfy large portions of the com-munity. But those who are dissatisfied with aparticular decision wil1 be more likely to ac-quiesce in it if they perceive that their views and

interests were given a fair hearing. If the deci-sion-maker has frankly laid the competing con-siderations on the table, so that the publicknows the worst as well as the best, he is unlike-

Z@D. L. Bazelon, ‘(coping With Technology Through th e Legal

Process, ” 62 Cornell Law Review 817,825, June 1977.

ly to find h imself accused of high-handedness,deceit, or cover-up. We simply cannot afford todeal with th ese vital issues in a manner that in-vites public cynicism and distrust.

The manner in which the Guidelines them-selves evolved has been controversial. (For adetailed d iscussion see app. III-A.) Initially, thescope and nature of the problem was defined bythe scientific comm unity; NIH organized RACalong the lines suggested by the N AS comm itteeletter referred to in app. III-A. One of the goalsof RAC was to recomm end guidelines for rDNAexperiments; it was not charged with consider-ing broader ethical or p olicy issues or th e fund a-mental question of whether the research shouldhave been p ermitted at all. The original Guide-lines were produced by a committee havingonly one nonscientist.

In late 1978, the Secretary of HEW signif-

icantly restructured RAC and modified theGuidelines in order to increase the system’saccountability to the pu blic, to “provide th e op-portunity for those concerned to raise anyethical issues posed by recombinant DNA re-search”{ and to m ake RAC “the pr incipal ad -visory body . . . on recombinant DNA policy. ”

27

However, it has remained in large part a tech-nically oriented body. Its charter was notchanged in this respect; the Guidelines them-selves state that its advice is “primarily scientificand technical, ” and matters presented for itsconsideration h ave continued to be mostly tech-nical. One area where RAC has played a signifi-cant policy role, however, is in dealing with theissue of voluntary compliance by ind ustry.

It could be argued th at the system did p rovidefor sufficient pu blic inpu t into th e formulationof the problem* and that no other formulationwas realistic. The two meetings in 1976 and1977 of the NIH Director’s Advisory Committeeand the hearing chaired by the general counselof HEW in the fall of 1978 prov ided the op por-tunity for public comment on the overall Fed-

ZTJoSeph A. Ca]ifano, “Notice of Revised Guidelines—Recombi-

nant DNA Research, ” 43 F.R. 60080-60081, Dec. 22, 1978.q The problem w as conceived in terms of how to permit the e.

search to be done while limiting th e physical risks to an acceptablelevel. Other formulations were p ossible, the broadest b eing howto limit al l risks, including cultural ones, to an acceptable level.Such a formulation could have resulted in a prohibition of theresearch.



Ch. 11-Regulation of Genetic Engineering q 22 3

One conflict of interest not solved by expand-ing the diversity of the RAC’s membership is in-stitutional in nature. N IH, the agency having p ri-mary responsibility for developing and adm inis-tering the Guidelines, views its mission as one of promoting biomedical research. Although the

Guidelines are not regulations, they containmany of the elements of regulations. They setstand ards, offer a limited means to mon itor forcompliance, and provide for enforcement, atleast for institutions receiving NIH grants to dorDNA work; thus, they may be consideredquasi-regulatory. Regulation is not only foreignbut antithetical to NIH’s mission. The currentDirector stated publicly at the June 1980 RACmeeting that the role of NIH is not one of aregulator, a role that must be avoided. Underthese circumstances, perhaps another agency,or another part of DHHS, might be more appro-

priate for overseeing the Guidelines, since theattitud es and priorities of promoters are u suallyquite different from that of regulators.

If RAC has always been essentially a technicaladvisory body, who then h as mad e the value de-cisions concerning the acceptability of the riskspresented by rDNA and the means for dealingwith them? The final decisionmaker has beenthe Director of NIH, w ith the notable exceptionin the case of the 1978 Guidelines, which con-tained the significant procedural revisionsneeded to m eet Secretary Califano’s appr oval.

31

The Director did have access to diverse pointsof view th rough the Director’s Advisory Com-mittee meetings and the public hearings heldbefore the 1978 Guidelines. (See app. III-A.) Inaddition, major actions were always accom-panied by a statement discussing the relevantissues and explaining the basis for the decisions;after the 1978 revisions, major actions had to beproposed for public comment before decisionswere m ade. In theory, it may have been p refer-able for the public to have been substantially in-volved in the actual formation of the originalGuidelines rather than simply to have reacted toa finished product. However, this probablywou ld have slowed the process at a time whenthe, strong d esire of the m olecular biologists to

3’Califano, op. cit.

q

use the rDNA techniques could have threatenedthe notion of self-regulation. Today, there ap-pears to be reasonable opportunity for publicinput through the process of commenting onprop osed actions.

Conclus ion

The Guidelines are the result of an extraor-dinary, conscientious effort by a combination of scientists, the public, and the Federal Govern-ment, all operating in an unfamiliar realm. Theyappear to be a reasonable solution to the prob-lem of how to m inimize the risks to health andthe environment posed by rDNA research in anacademic setting, while permitting as much of that research as possible to proceed. They donot in any way deal with other molecular genet-ic techniques or with the long-term social or

ph ilosophical issues that m ay be associated w ithgenetic engineering.

The Guidelines have been an evolving docu-ment. As more has been learned about rDNAand molecular genetics, containment levelshave been significantly lowered. Also, the de-gree of Federal oversight has been substantiallylessened. Und er the Novem ber 1980 Guidelines,virtually all responsibility for m onitoring com-pliance is placed on the IBCs. NIH’s role will in-volve prim arily: 1) continuing interp retation of the Guidelines, 2) certifying new host-vector

systems, 3) serving as a clearinghouse of infor-mation, 4) continuing risk assessment experi-ments, and 5) coordinating Federal an d local ac-tivities.

The most significant short-term limitation of the Guidelines is the way they deal with com-mercial applications and prod ucts of rDNA tech-niques. Although large-scale containment levelsand related administrative procedures exist,there are several reasons for questioning the ef-fectiveness of the voluntary compliance con-cept. The most serious problem has been thelack of expertise in fermentation technology onRAC. In addition, since the Gu idelines are notlegally bind ing up on indu stry, the NIH lacks en-forcement authority, although there has beenno evidence of industrial noncompliance. Final-ly, because of its role as a promoter of bio-




med ical research, NIH cannot be expected to act procedural safeguards designed to accommo-aggressively to fill this regulatory void. date social values and to limit conflicts of in-

As a model for societal decisionmaking onterest. The only major criticism is that proce-dural safeguards and public input were not sig-

technological risks, the system created by theGuidelines could serve as a valuable precedent.

nificant factors when the rDNA problem wasfirst add ressed.

It does a reasonable job of combining substan -tive scientific evaluation of technical issues with

o t h e r m e a n s o f r e g u l a t i o n

There are three other means available forregulating molecular genetic techniques andtheir products—current Federal statutes, tortlaw and. workmen’s compensation, and Stateand local laws. These all may be used to remed ysome of the limitations of the Guid elines.

Fede ra l s ta tu t e s

The question of whether existing Federalstatutes provide adequ ate regulatory auth orityfirst arose with respect to rDNA research. InMarch 1977, the Interagency Committee con-cluded that while a number of statutes* couldprov ide au thority to regu late specific ph ases of work with rDNA, no single one or combinationwou ld clearly reach all rDNA r esearch to th e ex-tent deemed necessary by the Committee. Fur-thermore, while some could be broadly inter-

preted , the Committee believed that regu latoryaction taken on the basis of those interpreta-tions w ould be subject to legal challenge.

32This

was the basis for th eir conclusion th at sp ecificlegislation w as needed and was one of the rea-sons behind the legislative effort discussed inapp. III-A.

With respect to commercial uses and prod-ucts of rDNA and other genetic techniques, amuch more certain basis for regulation exists.Many of the Federal environmental, productsafety, and public health laws are directedtowar d ind ustrial processes and p rodu cts. To a

“1’he Committee concentrated on the following statutes: 1) the

Occupational %fety and Health Act (29 IJ.S.C. S651 et. seq.); 2) theToxic Sul]stiin~es Control Act (15 IJ.S.(:, $2601 et. seq.); 3) the Haz -ardousMiiterials ‘1’ransportation Act (49 LJ. S.C. ~ 1801 et, seq.); and4) sec. 361 of th e Public Health Service Act (42 LJ.S.C. $264).

az/nterjm Re/lort Of the Federal Interagency CQnlmittee on Recom -

binant DNA Research: Suggested Elemenfsfor f.e.gislation, op. cit.

large extent, the genetic technologies w ill pro-duce chemicals, foods, and drugs—as well aspollutant byproducts—that will clearly comewithin the scope of these laws. ’However, theremay be limitations in these laws and questionsof their interpretation that may arise with re-

spect to the manufacturing process, whichemploys large quantities of organisms, andwhen there is an intentional release of micro-organisms into the environment—e.g., for clean-ing up pollution. For a list of pertinent laws, seetable 36.)

The Federal Food, Drug, and Cosmetic Act(FEDCA) and section 351 of the Public HealthService Act (42 U.S.C. 262) give FDA authorityover foods, dru gs, biological prod ucts (such asvaccines), med ical d evices, and veterinary m edi-cines. This authority will also apply to thoseprodu cts wh en they are mad e by genetic engi-

Table 36.—Statutes That Will Be Most Applicableto Commercial Genetic Engineering

1. Federal Food, Drug, and Cosmetic Act (21 U.S.C. $301et. seq.)

2. Occupational Safety and Health Act (29 U.S.C. $651 et.seq.)

3. Toxic Substances Control Act (15 U.S.C. $2601 et.seq.)

4. Marine Protection, Research, and Sanctuaries Act (33U.S.C. $1401 et. seq.)

5. Federal Water Pollution Control Act, as amended bythe Clean Water Act of 1977 (33 U.S.C. $1251 et. seq.)

6. The Clean Air Act (42 U.S.C. $7401 et. seq.)7. Hazardous Materials Transportation Act (49 U.S.C.

$1801 et. seq.)8. Solid Waste Disposal Act, as amended by theResource Conservation and Recovery Act of 1976 (42U.S.C. $6901 et. seq.)

9. Public Health Service Act (42 U.S.C. $301 et. seq.)10. Federal Insecticide, Fungicide, and Rodenticide Act (7

U.S.C. $136 et. seq.)





neering methods. However, interpretive ques-

tions arising out of the unique nature of the

technologies—such as the type of data nec-

essary to show the safety and efficacy of a new

drug produced by rDNA techniques–will have

to be resolved by the administrative process on

a case-by-case basis.

FDA has not published any statements of of-

ficial policy toward products made by genetic

engineering. Since it has different statutory au-thority for different types of products, it is like-

ly that regulation will be on a product-by-prod-

uct basis through the appropriate FDA bureau.

Substances produced by genetic engineering

will generally be treated as analogous products

produced by conventional techniques with re-

spect to standards for chemistry, pharmacolo-

gy, and clinical protocols; however, quality con-

trols may have to be modified to assure continu-ous control of product purity and identity. In

addit ion, for the t ime being, the Bureau of

Drugs and the Bureau of Biologics will require a

new Notice of Claimed Investigational Exemp-

tion for a New Drug and a new N ew Drug Appli-

cation for products made by rDNA technology,

even if identity with the natural substance or

with a previously approved drug is shown. This

policy is based on the position that drugs or

biologics made by rDNA techniques have not

become generally recognized by experts as safe

and effective and therefore meet the statutory

defin ition of a “new d rug. ’’33

*

FFDCA also permits regulation of drug, food,

and device manufacturing. Certain FDA regula-

tions, called Good Manufacturing Practices, are

designed to assure the quality of these produ cts.

FDA may have to revise these to accommodate

genetic technologies; it has the authority to doso. It probably does not have the authority to

use these regulations to address any risks to

workers, the public, or the environment, since

FFDCA is designed to protect the consumer of

the regulated product.

ssMinUteS of the In dustrial Practices Sub committee of the Fed-

eral Interagency Advisory Committee on Recombinan t DNA Re-search, Dec. 16, 1980, p. 3.

*Sec. 201(p) of th e FFLX:A (21 IJ.S.(:. $321(p)) defin es a new drLIg

as “any drug . . . the comp osition d ’ which is such that such drLJg

is not .gen(?rtil]y re(mgnized, i]n](~[l~ t?xperts (Iuiilitied by scient i f ic

tl’ilillill~ and experience . . iIS Silf[? illld Pt”t”(?d i~’e . .”

The statute most applicable to worker health

and safety is the Occupational Safety and Health

Act, which grants the Secretary of Labor broad

power to require employers to provide a safe

wor kplace for their emp loyees. This pow er in -

cludes the abil i ty to require an employer to

modify work practices and to instal l controltechnology. The statute creates a general duty

on employers to furnish their employees with aworkplace “free from recognized hazards that

are causing or are likely to cause death or seri-

ous ph ysical harm, ” and it requires emp loyers

to comply with occupational safety and health

standards set by the Secretary of Labor. Accord-

ing to a recent Supreme Court case, a standard

may be promulgated only on a determination

that it is “reasonably necessary and appropriate

to remedy a significant risk of material health

impairment. ”34

Because these fairly stringent re-

quirements l imi t the Act’s appl icabi l i ty torecognized hazards or significant r isks, the

statute could not be used to control manufactur-

ing where the genetic techniques presented on-

ly hypothetical risks. However, it should be ap-

plicable to large-scale processes using known

human toxins, pathogens, or their DNA.

The Secretary of Labor is also directed to ac-

count for the “urgency of the need” in es-

tablishing regulatory priori t ies. How the De-

partment of Labor will view genetic technol-ogies within its scale of priorities remains to be

seen. NIOSH, the research organization createdby this statute, has been stud ying rDNA produ c-

tion methods to determine what risks, if any,

are being faced by workers. It has conductedfact-f inding inspections of several manufac-turers, and it is planning a joint project with

EPA to assess the adequacy of current control

technology. In addition, a group established by

the Center for Disease Control (CDC) together

with NIOSH will be making recommendations

on: 1) the medical surveillance of potentially ex-

posed workers, 2) the central collection and

analysis of medical data for epidemiological pur-

poses, and 3) the establishment of an emergencyresponse team.

35

s4/ndustria/ Union Department, AFL-C1O v. American Rtrolellm

Institute, 100 S. Ct . 2844,2863, 1980.3SMinute~ of th e [ndustria] practices Subcommittee of th eFedera l

Interagency Advisory Committee on Recombinant DNA Research,Dec. 16, 1980, op. cit., p , 6.




tion, Research, and Sanctuaries Act prohibitsocean du mp ing without an EPA permit of anymaterial that wou ld “un reasonably degrade orendanger human health, welfare, or amenities,or the m arine environm ent, ecological systems,or economic potentialities. ”

39“Material” is de-

fined as “matter of any kind or d escription, in-cluding. . . biological and laboratory waste. . . and indu strial . . . and other waste. ”

40The

Federal Water Pollution Control Act regulatesthe d ischarge of pollutant s (which includ e bio-logical materials) into U.S. waters, and the SolidWaste Disposal Act regulates hazardou s w astes.The Clean Air Act regulates the discharge of airpollutants, which includes biological materials.Especially applicable is section 112 (42 U.S.C.7412), which allow s EPA to set emission stand -ards for hazardous air pollutants—those forwh ich standard s have not been set und er other

sections of the Act and wh ich “m ay reasonablybe anticipated to result in an increase in m ortali-ty or an increase in serious irreversible, or in-capacitating reversible, illness. ” The HazardousMaterials Transportation Act covers the inter-state transportation of dangerous articles, in-cluding etiologic (disease-causing) agents. TheSecretary of Transportation may designate ashazardous any material that he finds “may posean unreasonable risk to health and safety orproperty” w hen transported in commerce in aparticular quantity and form.

41

Section 361 of the Public Health Service Act(42 U.S.C. $264) authorizes the Secretary of HEW (now DHHS) to “. . . make and enforcesuch regulations as in his judgment are neces-sary to prevent the introduction, transmission,or spread of communicable diseases . . . .“ Be-cause of the broad discretion g iven to th e Sec-retary, it has been argu ed that this section pro-vides sufficient authority to control all rDNA ac-tivities. * Others have argued that its purpose isto protect only human health; for regulations tobe valid, there wou ld have to be a supp ortablefinding of a connection between rDNA and

3933 U.s.c. $1412.

to 33 U.S.CO ~ 1402(c).4149 U.S.C. ~ 1803.

“On Nov. 11, 1976, the Na tural Resou rces Defense Cou ncil andthe Environmental Defense Fund petitioned the Secretary of HEWto promu lgate regulations concerning DNA und er this Act.

hu man disease. In any event, HEW declined toprom ulgate any regulations.

The following conclusions can therefore bemade on the applicability of existing statutes.First, the products of genetic technologies—such as drugs, chemicals, pesticides,** and

foods—would clearly be covered by statutesalready covering these generic categories of materials. Second, uncertainty exists for regu-lating either production methods using en-gineered micro-organisms or their intentionalrelease into the environment, when risk has notbeen clearly demonstrated. Third, the regu-latory agencies have begun to study the situa-tion but, have not promulgated specific regu-lations. Fourth, since regulation will be dis-persed throughout several agencies, there maybe conflicting interpretations unless active ef-forts are mad e by the Federal Interagency Com-mittee to develop a comp rehensive, coordinatedapproach.

T or t law a nd w ork m e n’s compe nsa t ion

Statutes and r egulations are usu ally directedat preventing certain types of conduct. Whiletort law strives for the same goal, its primarypurpose is to compensate injuries. (A tort is acivil wrong, other than breach of contract, forwhich a court awards damages or other relief. )By its nature, tort law is quite flexible, since ithas been d eveloped primar ily by the courts on acase-by-case basis. Its basic principles can easilybe applied to cases where injuries have beencaused by a genetically engineered organism,prod uct, or p rocess. It therefore can be app liedto cases involving genetic technologies as ameans of compensating injuries and as an incen-tive for safety-conscious conduct. The most ap-plicable concepts of tort law are negligence andstrict liability. (A related body of law—work-men’s compensation—is also p ertinent. )

Negligence is defined as conduct (an act or anomission) that involves an un reasonable risk of

harm to another person. For the injured partyto be compensated, he must prove in court that:1) the defendant’s conduct was negligent, 2) the

q q Pesticides iire sut}ject to the ti’ederal Insecticide Fungicide,i ind Rodt)tlt icidr A(Y, 7 L r. S.(; ~ 136 et . S(>(l..




defendant’s actions in fact caused the injury,and 3) the injury was not one for which com-pensation should be denied or limited becauseof overriding policy reasons.

Because of the newness of genetic technol-ogy, legal stand ards of cond uct (e.g., what con-

stitutes unreasonable risk) have not been ar-ticulated by the courts. If a case were to ar ise, acourt would undoubtedly look first to theGuidelines. Even if a technique other th an rDN Awere involved, they would provide a generalconceptual framework for good laboratory andindustrial techniques. Other sources for stand-ards of conduct include: 1) CDC’s guidelines forworking with hazardous agents; 2) specific Fed-eral laws or regulations, such as those und er thePublic Health Service Act covering the inter-state transportation of biologic products andetiologic agents; and 3) industrial or profes-

sional codes or customary practices, such asgenerally accepted containment practices in thepharmaceutical industry or in a microbiologylaboratory. Compliance with these standards,however, does not foreclose a finding of neg-ligence, since the courts make the ultimate judg-ment of what constitutes proper conduct. Inseveral cases, courts h ave d ecided that an entireindustry or profession has lagged behind thelevel of safe practices demanded by society. *Conversely, noncompliance with existing stand-ards almost surely will result in a finding of negligence, if the other elements are also p res-ent.

Causation m ay be d ifficult to prove in a caseinvolving a genetically engineered product ororganism. In the case of injury caused by a path-ogenic micro-organism —e.g., it may be difficultto isolate and identify the micro-organism an dvirtually impossible to trace its origin, especiallyif it had only established a transitory ecologicalniche. In addition, it might be difficult toreconstruct the original situation to determineif the micro-organism simply escaped despite

‘For example, see: The T. J. Hooper, 60 F. 2d 737 (2d Cir. 1932),concerning tugboats; and He/hhg v. Carey, 519 P. 2d 981 (1974),where the court held that the general practice among oph thalmo-logists of not p erforming glaucoma tests on symptomatic patientsun der 40 [because th ey had on ly a one in 25,000 chance of havin gthe disease) would not prevent a finding of negligence when su cha patient developed the d isease.

precautions or if culpable human action was in-volved. On th e other han d, if a micro-organismor toxin is identified, it may be so uniquebecause of its engineering that it can be read ilyassociated with a company known to produce itor with a scientist known to be working withit. * *

The law recognizes that not every negligentact or omission that causes harm should resultin liability and compensation—e.g., the conceptof “foreseeable” harm serves to limit a de-fendant’s liability. The underlying social policyis that the d efend ant shou ld not be liable for in-

juries so random or unlikely as to be not rea-sonably foreseeable. This determination is madeby the court. In the case of a genetically en-gineered organism, extensive harm would prob-ably be foreseeable because of the organism’sability to reprodu ce; how that harm could occur

might not be foreseeable.

Unlike negligence, strict liability does not re-quire a finding that the defendant breachedsome du ty of care owed to th e injured person;the fact that the injury was caused by the de-fendant’s cond uct is enough to imp ose liabilityregardless of how carefully the activity wasdone. For this doctrine to apply, the activitymust be characterized as “abnormally dan-gerous.” To determ ine this, a court wou ld look at the following six factors, no one of which isdeterminative:

42

1. existence of a high risk of harm,2. great gravity of the harm if it occurs,3. inability to eliminate the risk by exercising

reasonable care,

q “I f several compan ies were workin g with the micro-organism,it could b e impossible to prove which compan y produced the par-ticular ones that caused the h arm. A recent California SupremeCourt case, Sindell v. Abbott Laboratories, 26 CaL3d 588, 1980,

could provide a way around this problem if the new th eory of liability that it establishes becomes widely accepted by courts inother jurisdictions. The Court ruled that wom en wh ose mothersh i i d taken diethy]stilbest rol, a dru g that allegedly caused cancer intheir daugh ters, could proceed to trial against manufacturers of the drug, even though most of the plaintiffs would not be ab le to

show wh ich particular manu facturers produced the drug. TheCourt said that when the defend ant manu facturers had a substan-tial share of the produ ct market, liability, if found, wou ld b e ap-

portioned among the defendan ts on the basis of their marketshare. A particular defendant could escape liability only byproving it could not have made the d rug.

dz~es.tatement &cond) of Torts $520 (1976).



Ch. 11—Regulation of Genetic Engineering . 229

4. extent to which the activity is not common,5. inappropriateness of the activity to the

place where it is done, and6. the activity’s value to th e commun ity.

Given the current consensus about the risksof genetic techniques, it would be difficult to

argue that t he d octrine of strict liability shou ldapp ly. However, in the extremely unlikely eventthat a serious, widespread injury does occur,that alone would probably support a court’s de-termination that the activity was abnormallydangerous, regardless of its probability. In suchcases, the courts have generally relied on theprinciple of “enterprise liability ’’–that those en-gaged in an enterprise should bear its costs, in-cluding the costs of injuries to others.

43

For either negligence or strict liability, theperson causing the harm is liable. Under the

legal principle of respondent superior, liability isalso imputed from the original actor to peopleor entities who have a special relationship w ithhim—e.g., employers. Thus, a corporation canbe liable for the torts of its scientists or prod uc-tion workers. Similarly, a university, an IBC, aBiological Safety Officer, and a PI would prob-ably be liable for the torts of scientists and stu-dents under their direction.

Another body of law designed to compensateinjuries deserves brief mention. Workmen’scompensation is a statutory scheme adopted by

the States and—for specific occupations or cir-cumstances—by the Federal Government tocompensate injuries without a need for show ingfault. The employee need only show that the in-

jury was job-related. He is then compensated bythe employer or the employer’s insurance com-pan y. It would clearly app ly to genetic engineer-ing.

Tort law and workmen’s compensation willbe available to compensate any injuries re-sulting from th e use of molecular genetic tech-niques, especially from th eir commercial appli-cation. Tort law may also indirectly preventpotent ially hazard ous actions, althou gh the de-

43R. DWOrkln, “giocatastrophe and the Law: Legal Aspects of Re-

combinant D NA Research, ” in The Recombinant DNA Debate,

Jackson and S titch (eds.) (Englewood Cliffs, N. J.: Prentice-Hal], Inc.1979), pp . 219, 223.

terrent effect of compensation is less efficientthan direct regulation—e.g., the threat of law-suits will not necessarily discourage high-risk activities where problems of proof make re-covery unlikely, where the harm may be smalland widesp read (as w ith mild illness suffered by

a large number of peop le), or where p rofits areless than the cost of prevention bu t greater thanexpected d amage aw ard s and legal costs.

Tort law has two other limitations. First, tortlitigation involves high costs to th e plaintiff, andindirectly to society. Second, it cannot adequate-ly compensate th e victims of a catastroph ic sit-uation where liability would bankrupt thedefendant.

S t a t e a n d l o ca l l a w

Under the 10th am endm ent to the Constitu-tion, all powers not delegated to the FederalGovernment are reserved for the States or thepeop le. One of those is the pow er of the Statesand mu nicipalities to protect the health, safety,and welfare of their citizens. Thus, they canregulate genetic engineering.

The reasons espoused in favor of local regula-tion are based on the traditional concept of localautonomy; those most likely to suffer anyadverse affects of genetic engineering shouldcontrol it. Also, local and State governments areusua lly m ore accessible to pu blic input than the

Federal Government. Consequently, judgmentson the acceptability of the risks will moreprecisely reflect the will of the segment of thepublic most directly affected.

A number of arguments have been madeagainst local as opposed to Federal regulation.The primary one is that regulation by States andcommu nities would give rise to a random p atch-work of confusing and conflicting controls. Inaddition, States and especially localities may nothave the same access as the Federal Govern-ment to the expertise that should be used in the

formulation of rational controls. Finally, anyrisks associated with rDNA or other techniquesare not limited by geographic boundaries;therefore, they ought to be dealt with national-ly. The above arguments reflect the positionthat r egulation of genetic technologies is a na-

76-565 0 - 81 - 16




tional issue that can be handled most effectivelyat the Feder al level.

A few jurisdictions have u sed their auth orityin the case of rDNA, * The most comprehensiveregulation was created by the States of Mary-

q Cambridge, Mass., established a citizens’ study group that rec.

ommend ed that researchers be subject to some add itional re .striations beyond those of the Guid elines. These were embodied inan ordinan ce passed by the City Council on Feb. 7, 1977. Serkeley,Calif., passed an ordinance requiring p rivate research to conformto the Gu idelines. Similar ordinances or resolutions were passedby Prin ceton, N.J., Amh erst, Mass., and Emeryville, Calif.

land and New York.4445 Cur rently, ther e is little,

if any, effort on the State or local level to passlaws or ordinances covering rDNA or similargenetic techniques, and there is little activityund er the existing laws.

4aAnnotated Code of Maryland, art. 43 ~~ 898-910 (SUpp. 19781.

45Mc~jnney’s C“onsoljdated Laws of fVew York, Public Health ~w ,

art. 32-A S$3220-3223 (supp. 1980)

Conc lus ion

The initial question with respect to regulatinggenetic engineering is how to define the scopeof the problem. This will depend largely on

what groups are involved in that p rocess andhow they view the nature, magnitude, andacceptability of the risks. Similarly, the meansof addressing the problem will be determinedby how it is defined and wh o is involved in theactual decisionmaking process. For these rea-sons, it is important that regu latory m echanismscombine scientific expertise w ith p rocedu res toaccommodate the values of those bearing therisk so that society may have confidence inthose mechanisms.

Currently, genetic techniques and their prod-

ucts are regulated by a combination of the

Guidelines, Federal statutes protecting healthand the environm ent, some State or local laws,and the judicially created law of torts, wh ich is

available to compensate injuries after they oc-cur. In most cases, this system ap pears ad equateto deal with the risks to health and the environ-ment. However, there is some concern regard-ing commercial applications for the followingreasons: 1) the voluntary applicability of theGuidelines to ind ustry , 2) RAC’s insu fficient ex-pertise in fermentation technology, 3) the po-tential interpretive problems in applying ex-isting law to the workplace and to situationswhere micro-organisms are intentionally re-leased into the enviornment, and 4) the absenceof a definitive regulatory posture by the

agencies.

I s s u e a n d O p t i o n s

ISSUE: H o w c o u l d C o n g r e s s a d d r e s s t h er i s k s a s s o c i a t e d w i t h g e n e t i c e n -g i n e e r i n g ?

A number of options are available, rangingfrom deregulation through comprehensive newregulation. An underlying issue for most of

these options is: What are the constitutionalconstraints placed on congressional regu lationof molecular genetic techniques, particularlywhen they are used in research? (This is dis-cussed in ap p. III-B.)

OP TIONS :

A: Congress could maintain the status quo by let- ting NIH and the regulatory agencies set the Federal policy.

This option requires Congress to determinethat legislation to remedy the limitations in cur-

rent Federal oversight would result in unneces-sary and burdensome regulation. No knownharm to health or the environment has oc-curred un der the current system, and the agen-cies generally have significant legal authority




and expertise that should perm it them to adaptto most new problems posed by genetic engi-neering. The agencies have been consultingwith each other through the Interagency Com-mittee, and the th ree agencies that w ill play themost important role in regulating large-scale

commercial activities—FDA, OSHA, and EPA—have been studying the situation.

The disadv antages of this option are the lack of a centralized, uniform Federal response tothe problem, and the possibility that risksassociated w ith commercial app lications w ill notbe adequately addressed. Certain applications,such as the use of micro-organisms for oil re-covery are not unequ ivocally regulated by cur-rent statut es; broad interp retations of statutorylanguage in order to reach these situations maybe overturned in court. Conflicting or redun-dant regulations of different agencies wouldresult in unnecessary burdens on those regu-lated. In ad dition, some comm ercial activity isnow at the pilot plant stage, but the responsibleagencies have yet to establish official policy andto devise a coordinated p lan of action.

B: Congress could require that the Federal Inter- agency Advisory Committee on Recombinant DNA Research prepare a comprehensive report on its members’ collective authority to regulate rDNA and their regulatory intentions.

The Industrial Practices Subcommittee of thisCommittee has been studying agency authorityover commercial rDNA activities. Presently,there is little official guidan ce on r egulatory re-quirements for companies that may soon mar-ket products made by rDNA methods. -e.g.,companies are building fermentation plantswithout know ing what d esign or other require-ments OSHA may m andate for worker safety.As was stated by former OSHA head, Dr. EulaBingh am, it will take at least 2 years for OSHA toset standards, if the current NIOSH study showsa need for them.

46

A congressionally mandated report would

assure fu ll consideration of these issues by th eagencies and expedite the process. It could in-

46[,etter from Dr, ~llla Bingham, Assistant %?cretary fo r ~ccupa-

tional Safety and H ealth, to Dr. Don ald Fredrickson, Director,N IH ,Sept. 24, 1980.

elude the following: 1) a section prepared byeach agency that assesses its statutory authorityand articulates what activities and products willbe considered to come w ithin its jurisd iction, 2)a sum mary section th at evaluates the adequacyof existing Federal statutes and regulations as a

whole with respect to commercial genetic en-gineering, and 3) a section pr oposing an y specif-ic legislation considered to be necessary.

The principal disadvantages of this option arethat it maybe u nnecessary and imp ractical. Theagencies are studying the situation, which mustbe done before they can act. Also, it is ofteneasier and more efficient to act on each case asit arises, rather than on a hypothetical basisbefore the fact.

C: Congress could require Federal monitoring of all rDNA activity for a limited number o f

years.

This option rep resents a “wa it and see” posi-tion by Congress and the middle ground be-tween th e status quo an d full regulation. It rec-ognizes and balances the following factors: 1)

the absence of demonstrated harm to humanhealth or the environment from genetic en-gineering; 2) the continuing concern th at genet-ic engineering presents risks; 3) the lack of suf-ficient knowledge from which to make a final

judgment; 4) the existence of an oversight mech-anism that seems to be working well, but that

has clear limitations w ith respect to commercialactivities; 5) the virtual abolition of Federalmonitoring of rDNA activities by the recentamendments to the Guidelines; and 6) the ex-pected increase in commercial genetic engineer-ing activities.

Monitoring involves the collection an d eval-uation of information about an activity in orderto know what is occurring, to determine theneed for other action, and to be able to act if necessary. More specifically, this option wouldpr ovide a da ta base that could be used for: 1) de-termining the effectiveness of voluntary compli-

ance with the Guidelines by industry and man-datory compliance by Federal grantees, 2) de-termining the qu ality and consistency of IBC de-cisions and other actions, 3) continu ing a form alrisk assessment program, 4) identifying vague



Ch. 11-Regulation of Genetic Engineering q 23 3

in fermentation and other industrial technolo-gies to RAC if production, as well as research, isto be adequately covered. In addition, the rec-ommendations for large-scale containment pro-cedures would have to be made part of theGuidelines.

The major changes would have to be madewith respect to enforcement. Present penaltiesfor noncompliance—suspension or terminationof research funds—are obviously inapplicable toindustry. In addition, procedures for monitor-ing compliance could be strengthened. Some of the elements of option C could be used. Anadd ed or alternative approach wou ld be to in-spect facilities.

The main disadvantage of this option is thatNIH is not a regu latory agency. Since NIH hastraditionally viewed its mission as promoting

biomedical research, it w ould have a conflict of interest between regulation and promotion.One of the regulatory agencies could be giventhe authority to enforce the Guidelines and toadopt changes therein. NIH could then continuein a scientific adv isory role.

E. Congress could require an environmental impact statement and agency approval before any genetically engineered organism is intentionally released into the environment.

There have been numerous cases where ananimal or p lant species has been introdu ced intoa new environment and h as spread in an uncon-trolled and undesirable fashion. One of theearly fears about rDNA was that a new path-ogenic or otherwise undesirable micro-orga-nism could establish an environmental niche.Yet in pollution control, mineral leaching, andenhan ced oil recovery, it might be desirable torelease large numbers of engineered micro-or-ganisms into the environment.

The Guidelines currently prohibit deliberaterelease of any organism containing rDNA with-out ap proval by the Director of NIH on advice of

RAC. The obvious d isadvantage of this prohibi-tion is th at it lacks the force of law. The releaseof such an organism without NIH approvalwould be a prima facie case of negligence, if theorganism caused harm. However, it may bemore desirable social policy to attempt to pre-

vent this type of harm through regulationrather than to compensate for injuries throughlawsu its. Another p ossible disadva ntage of thepresent system is that app roval maybe grantedon a finding that the release would present “nosignificant risk to health or the environment;” a

tougher or more specific standard than this maybe d esirable.

A required study of the possible consequen-ces following the release of a geneticallyengineered organism, especially a micro-orga-nism, would be an important step in ensuringsafety. This option could be implemented by re-quiring those p roposing to release the organismto file an impact statement with an agency suchas NIH or EPA, wh ich w ould then gran t or denypermission to release the organism. A disad-vantage of this option is that companies and in-

dividu als might be discouraged from developinguseful organisms if this process became tooburdensome and costly.

F. Congress could pass legislation regulating all types and phases of genetic engineering from research through commercial production.

The main ad vantage of this option would beto deal comprehensively and directly with therisks of novel molecular genetic techniques,rather than relying on the current patchwork system. A specific statute would eliminate theuncertainties over the extent to which present

law covers particular applications of genetic en-gineering, such as pollution control, and anyconcerns about the effectiveness of voluntarycompliance w ith the Gu idelines.

Other molecular genetic techniques, whilenot as widely used and effective as rDNA, raisesimilar concerns. Of the current techniques, cellfusion is the prime cand idate for being treatedlike rDNA in any regulatory framework. It per-mits the recombination of chromosomes of species that do not recombine naturally, and itmay perm it the DNA of latent viruses in the cells

to recombine into harmful viruses. No risk as-sessment of this technique has been done, andno Federa l oversight exists.

The principal arguments against this optionare that the current system appears to be work-ing fairly w ell, and that th e limited risks of the




techniques may not warrant the significantly in-creased regulatory burden and costs that wouldresult from such legislation. Congress will haveto decide if that system will remain adequate ascommercial activity grows.

If Congress were to decide on this option, thelegislation could incorporate some or all of op-tions C, D, and E. The present mechanismcreated by the Guidelines could be appropriate-ly modified to provide the regulatory frame-work. The m odifications could include a regis-tration and licensing system to provide infor-mation on wh at work w as actually being doneand a means for continuous oversight. Oneimportant typ e of information wou ld be h ealthand safety statistics gathered by monitoringworkers involved in the production of productsfrom genetically engineered orga nisms. Anoth-

er modification could be a sliding scale of penalties for violations, ranging from monetaryfines through revocation of operating licensesto criminal penalties for extreme cases.

It would not be necessary to create a newagency, wh ich w ould d up licate some of the re-sponsibilities of existing agencies. Instead, Con-gress could give these agencies clear regulatoryauthority by amending the appropriate statutes.Designating a lead agency would assure a moreuniform interpretation and application of thelaws.

G. Congress could require NIH to rescind theGuidelines.

This option requires Congress to determinethat th e risks of rDNA techniques are so insig-nificant that no control or oversight is nec-essary. Deregulation would have the advantageof allowing funds and personnel currently in-volved in implementing the Guidelines at theFederal and local levels to be u sed for other p ur-poses. In fiscal year 1980, NIH spent approxi-mately $500,000 in administering the Guide-lines; figures are not available for the analogouscost to academia and industry. Personnel hours

spent have not been estimated. Very few peoplework full-time on administering or complyingwith the Guidelines. NIH em ploys only six peo-ple full-time for this pu rpose, and som e institu-tions em ploy full-time biological safety person -nel. However, over 1,000 people nationally

devote some effor t to implementing theGuidelines—members of the IBCs and the scien-tists conducting the rDNA experiments whomu st take necessary steps to comp ly.

There are several reasons for retaining theGuidelines. First, sufficient scientific concernabout risks exists for the Gu idelines to proh ibitcertain experiments and require containmentfor others. Second, they are not particularlyburdensome, since an estimated 80 to 85 per-cent of all experiments can be done at thelowest containment levels and an estimated 97

percent will not require NIH approval. Third,NIH w ill continue to serve an important role incontinuing risk assessments, in evaluating newhost-vector systems, in collecting and dispersinginformation, and in interpreting the Guidelines.Fourth, if the Guidelines were abolished, regu -latory activity at the State and local levels couldagain become active. Finally, the oversight sys-tem has been flexible enough in the past to lib-eralize restrictions as evidence ind icated lowerrisk.

H. Congress could consider the need for

regulating work with all hazardous micro- organisms and viruses, whether or not they are genetically engineered.

Micro-organisms carrying rDNA, according toan increasingly accepted view, represent just asubset of micro-organisms and viruses, which,in general, pose risks. CDC has pu blished guide-lines for working w ith hazard ous agents such aspolio virus. However, such work is not cur-rently subject to legally enforceable Federal reg-ulations. It was not within the scope of thisstudy to examine this issue, but it is an em ergingone that Congress may w ish to consider.



C h a p t e r 12

P a t e n t i n g L i v i n g O r g a n i s m s

A l a n d m a r k d e c i s i o n

In a 5 to 4 decision (Diamond v. Chakrabarty,June 16, 1980), the Supreme Court ruled that amanmade mice-organism is patentable underthe current patent statutes. This decision wasalternately hailed as having “assured this coun-try’s technology futu re”

land denoun ced as cre-

ating “the Brave N ew World th at Aldous H uxleywar ned of. ”

2How ever, the Court clearly stated

that it was un dertaking only the narrow task of determining whether or not Congress, in enact-ing the patent statutes, had intended a man-made micro-organism to be excluded from pat-

entability solely because it was alive. Moreover,the opinion invited Congress to overturn thedecision if it disagreed with the Court’s inter-pretation.

‘Prepared Statement of C,enentech, Inc., cited in “Science MayPatent New Forms of Life, Justices Rule, 5 to 4,” Th e New York

‘rimes, Jun e 17, 1980, p. 1.‘Prepared statement of the Peoples’ Business Commission, cited

in “Science Ma y Patent N ew Forms of Life, Justices Rule, 5 to 4,”The New York Times, June 17, 1980, p. 1.

Congress may want to reconsider the issue of whether and to what extent it should specifi-cally provide for or prohibit the patentability of living organisms. While the judiciary operateson a case-by-case basis, Congress can considerall the issues related to p atentability at the sametime, gathering all relevant data an d taking tes-timony from the interested parties. The issuesinvolved go beyond the narrow ones of scien-tific capabilities and the legal interpretations of statutory wording. They require broader deci-sions based on pu blic policy and social values;

Congress has the constitutional authority tomake those decisions for society. It can act to re-solve the questions left unanswered by theCourt, overru le the decision, or develop a com-prehensive statutory approach, if necessary.Most importantly, Congress can draw lines; itcan specifically decide which organisms, if any,should be p atentable.

L e g a l p r o t e c t i o n o f i n v e n t i o n s

The inherent “right” of the originator of anew idea to that id ea is generally recognized, atleast to the extent of deserving credit for itwh en used by others. At the same time, it is alsobelieved that worthwhile ideas benefit societywh en th ey are widely av ailable. Similarly, whenan idea is embod ied in a tangible form, such asin a machine or industrial process, the inventorhas the “right” to its exclusive posession and usesimply by keeping it secret. However, if he maybe induced to disclose the invention’s details,

society benefits from the new ideas embodiedtherein, since others may build upon the newknowledge. The legal system has long recog-nized the competing interests of the inventorand the public, and has attempted to protect

both. The separate laws covering trade secretsand patents are the mean by w hich this is done.

Trade secre ts

The body of law governing trad e secrets rec-ognizes that harm has been done to on e personif another improperly obtains a trade secret andthen uses it personally or discloses it to others.A trad e secret is anything—device, formula, orinformation—which when used in a business

provides an advantage over competitors ig-norant of it—e.g., improper acquisition includesa breach of confidence, a breach of a specificpromise not to disclose, or an outright theft.Trade secrecy is derived from the common law,

237



Ch. 12—Patenting Living Organisms q 239

know n or used p ublicly by others in the UnitedStates, or 3) the invention was previously de-scribed in a U.S. or foreign patent or publica-tion. The inability to meet the n ovelty require-ment is another reason w hy p rodu cts of natureare unp atentable. Non obviousness refers to the

degree of difference between th e invention andthe prior art. If the invention would have beenobvious at the time it was m ade to a p erson withordinary skill in that field of technology, then itis not patentable. The policy behind the dualcriteria of novelty and nonobv iousness is that apatent should not take from the public some-thing which it already enjoys or potentiallyenjoys as an obvious extension of currentknowledge.

The final requirement—for adequate publicdisclosure of an invention—is know n as the en-ablement requirement. It is designed to ensurethat the public receives the full ,benefit of thenew knowledge in return for granting a limitedmonopoly. As a public document, the patentmu st contain a sufficiently d etailed descriptionof the invention so that others in that field of technology can build and use it. At the end of this description are the claims, which define theboundaries of the invention protected by thepatent.

The differences between trade secrets andpatents, therefore, center on the categories of inventions protected, the term and degree of

protection, and the disclosure required. Onlythose inventions meeting the statutory requ ire-ments outlined above qualify for patents andthen only for a limited time, whereas anythinggiving an advan tage over business competitorsqualifies as a trad e secret for an unlimited time.A patent requires full public disclosure, whiletrade secrecy requires an explicit and oftencostly effort to w ithhold information. The p at-ent law provides rights of exclusion againsteveryone, even subsequent independent inven-tors, while the trade secrecy law protects onlyagainst wrongful ap prop riation of the secret.

Any organism that both meets the broad defini-tion of a trade secret and may be lawfullyowned by a private person or entity can be pro-tected by that body of law, including micro-organisms, plants, animals, and insects. In ad di-tion, plants are covered specifically by two Fed-

eral statutes, the Plant Patent Act of 1930 andthe Plant Variety Protection Act of 1970. Fur-thermore, the Supreme Court has now ruledthat manmade micro-organisms are covered bythe patent statutes. Its determination of con-gressional intent in the Chakrabarty case wasbased significantly on an analysis of the twoplant protection statu tes.

Patent pr otection for plants w as not availableuntil Congress passed the Plant Patent Act of 1930, recognizing that not all plants were prod-ucts of nature because new varieties could becreated by m an. This Act covered new and dis-tinct asexually reproduced var ieties other thantuber-propagated p lants or those found in na-ture. * The requirement for asexual reproduc-tion was based on the belief that sexuallyreproduced varieties could not be reproducedtrue-to-type an d that it wou ld be senseless to tryto protect a variety that would change in thenext generation. To deal with the fact that or-ganisms rep rodu ce, the Act conferred the rightto exclude others from asexually reproducingthe p lant or from using or selling any plants soreproduced. It also liberalized the description

requirement for plants. Because of the impos-sibility of describing plants with the same de-gree of specificity as machines, their descriptionneed only be as comp lete as is “reasonably possi-ble.”

By 1970, plant breeding technology had ad-vanced to where new, stable, and uniform vari-eties could be sexually reprod uced. As a result,Congress provided patent-like protection tonovel varieties of plants that reproduced sexu-ally by passing the. Plant Variety Protection Actof 1970. Fungi, bacteria, and first-generationhybrids were excluded. * * Hybrids have a built-

L iv ing o r g a n i s m s

Although the law for p rotecting inventions isusually thought of as applying to inanimate ob-

jects, it also app lies to certain living organ isms.

q Approximately 4,500 plant paten ts have been issued to d ate,most for roses, apples, peaches, and chrysanthemums.

qq Originally, six vegetables—okra, celery, peppers, tomatoes,carrots, and cucumbers—were also excluded. On Dec. 22, 1980,

President Carter signed legislation (H.R. 999) amend ing the PlantVariety Protection Act to include these vegetables, to extend theterm of protection to 18 years, and to make certain technicalchanges.



Ch. 12—Patenting Living Organisms q 241

scope. Moreover, when these laws were last re-

modi f ied in 1952, the congress iona l commit tee

reports affi rmed the intent of congress that pat-

e n t a b l e sub j e c t ma t t e r “ i nc l ude a ny t h i ng unde r

the sun that is made by man. ”6The Court

acknowledged that not everything is patentable;

laws of nature, physical phenomena, andabstract ideas are not.

The Court found the Government’s argu-ments unpersuasive. Specifically, that passingthe Plant Patent A ct of 1930 and the Plant Varie-ty Protection Act of 1970, which excluded bac-teria, was evidence of congressional under-standing that section 101 did not apply to livingorganisms; otherwise; these statutes wouldhave been unnecessary. In disagreeing, theCourt stated th at the 1930 Act w as necessary toovercome the belief that even artificially bred

plants were unpatentable products of natureand to relax the written description require-ment, permitting a description as complete as is“reasonably possible. ” As for the 1970 Act, theCourt stated that it had been p assed to extendpatent-like p rotection to new sexually reprod uc-ing varieties, which, in 1930, were believed tobe incapable of reproducing in a stable, uniformmanner. The 1970 Act’s exclusion of bacteria,which indicated to the Government that Con-gress had not intended bacteria to be pat-entable, was considered insignificant for a num-ber of reasons.

The Government had also argued that Con-gress could not have intended section 101 tocover genetically eng ineered micro-organisms,since the technology was unforeseen at thetime. The majority responded that the very pur-pose of the patent law was to encourage new,un foreseen inventions, which was w hy section101 was so broadly worded. Furthermore, asfor the “gruesome parade of horribles”

7that

might possibly be associated w ith genetic engi-neering, the Court stated that the denial of apaten t on a m icro-organism might slow the sci-

entific work but certainly would not stop it; andthe consideration of su ch issues involves p olicy judgments that the legislative and executive

es , R@, N~ , , 1979, 82d ( : [ ] n g , , 2 d $ j ~ s s . , P. 5, 19,52; H . R. Rq ) t . No.

1 9 2 3 , m l IIong., 2 d Sess. j p . 6, 1 9 5 2 , c i ted in D i am~~~ v.

Chakrabarfy, 100” S. Ct. 2204, 2207 [ 1980).7Diamond t’. Chakrabarty, 100 S. Ct. 2204, 2211 (1980).

branches of Government, and not the courts,are competent to make. It further recognizedthat Congress could amend section 101 to spe-

cifically exclude genetically engineered orga-

nisms or could write a statute specifically de-

signed for them.

The dissenting Justices agreed that the issue

was one of statutory interpretation, but inter-

preted section 101 differently. They saw the

two plant protection Acts as strong evidence of

congressional intent that section 101 not cover

living organisms. In view of this, the dissenters

maintained that the majority opinion was ac-

tually extending the scope of the patent laws

beyond the limit set by Congress.

The stated narrowness of the Court’s decision

may limit its impact as precedent in subsequentcases th at raise similar issues, although not nec-essarily. Certainly, the decision applies to anygenetically engineered micro-organism. It is atechnical distinction without legal significancethat most of the work being d one on such orga-nisms involves recombinant DNA (rDNA) tech-niques, which Chakrabarty did not use. The realquestion is whether or not it would permit thepatenting of other genetically engineered or-ganisms, such as plants, animals, and insects.Any fears that the decision might serve as alegal precedent for the patenting of human be-ings in the distant future are totally groundless.

Under our legal system, the ownership of hu-mans is absolutely prohibited by the 13thamend ment to the Constitution.

Although the Chakrabarty case involved amicro-organism, there is no reason that its ra-tionale could not be applied to other organisms.In the majority’s view, th e crucial test for pat-entability concerned whether or not the micro-organism was manmade. Conceptually, there isnothing in this test that limits it to micro-organisms. The operative distinction is betweenhum anmad e and naturally occurring “things,”regardless of what they are. Thus, the Chakra- barty opinion could be read as precedent for in-cluding any genetically engineered organism(except humans) within the scope of section 101.Whether a court in a subsequ ent case will inter-

pret Chakrabarty broadly or narrowly cannot bepredicted.




Even if section 101 were interpreted as cover-ing other genetically engineered organisms,they probably could n ot be patented for failureto meet another requirement of the patentlaws—the enablement requ irement. It is gener-ally impossible to describe a living organ ism in

writing with enough detail so that it can bemad e on th e basis of that description. Relaxingthis requirement for plants was one reasonbehind the Plant Patent Act of 1930. For micro-organisms, the problem is solved by d epositing apublicly available culture with a recognized na-tional repository and referring to the accessionnum ber in the patent. * While such an ap proach

may be theoretically possible for animals and in-sects, it may be logistically impractical. How-ever, if tissue culture techniqu es advan ce to thepoint where genetically engineered organismscan be made from single cells and stored indefi-nitely in that form, there appears to be no rea-

son to treat them any differently than micro-organisms, in the absence of a specific statuteprohibiting their patentability.

q This procedure was accepted by the Court of Customs and Pat-

ent Appeals (CCPA) in uph olding a patent on a process using micro-organisms. Application of Argoudelis,” 434 F.Zd 1390 (CCPA 1970).This procedure should also be acceptable for patents on micro-organisms themselves.

P o t e n t i a l i m p a c t s o f t h e d e c i s i o n a n d

r e l a t e d p o l i c y i s su e s

During the 8-year history of the Chakrabartycase and the surrounding public debate, nu-merous assertions were mad e about the poten-tial impacts of permitting patents on geneticallyengineered organisms. They ranged from m oreimmediate effects on the biotechnology indus-

try, the patent system, and academic researchto the long-term impacts on genetic diversityand the food su pp ly. In add ition, two major pol-icy issues that hav e been raised a re the m oralityof patenting living organisms; and the prop rietyof permitting private ownership of inventionsfrom publicly funded research.

I mpac t s on indus t ry

The basic question for ind ustry is the extentto which permitting patents on genetically en-

gineered organisms w ill stimu late both their d e-velopment and the growth of the industries em-ploying th em. To ascertain this requ ires first anexamination of the theory and social policiesund erlying the patent system.

THE RELATIONSHIP BETWEEN PATENTSAND INNOVATION

The patent system is supposed to stimu late in-novation—the process by which an invention isbrought into commercial use—because the in-ventor d oes not receive financial reward s un til

the invention is used commercially. The Con-stitution itself presum es this, as do th e statutesenacted pursuant to the patent clause in articleI, section 8. Attempts have been made to subjectthis presumption to empirical analysis; but in-novation is extraordinarily complex and in-volves interacting factors that are difficult toseparate. In addition, the existence of patentsand trade secrets as alternative means for pro-tection m akes it almost impossible to study theeffects of patents alone on invention and in-novation. *

q A majo r reuson for the lack of empirical studies h as been th elack of appropriate data. The information available on the n umberof patents applied for and issued does not in dicate the importance,economic b enefits, or economic costs of inventions (whether p at-ented or u npatented) that may n ot have existed at all or may havebeen created more slowly if not for the patent system.S In Presi-




and not worth the time and money to prosecuteReliance on trade secrecy might th en extend itscomm ercial life.

Most compan ies wou ld p atent truly pioneerinventions, which often pr ovide the opp ortunityfor developing large markets. Moreover, pat-ents of this sort tend to have long commerciallives, since it is difficult to circumvent a p ioneerinvention and since any improvements are stillsubject to the pioneer patent. Furthermore, in-fringement is easy to detect because of the in-vention’s trailblazing nature.

The last two factors involve considerationssecondary to a product and its market. Ob-viously, any publication of the experimentsleading to an invention foreclose the option of trade secrecy. Also, company must evaluate theoptions of protection via either patenting or

trade secrecy in terms of their respective costeffectiveness.

IMPACT OF THE COURT’S DECISIONON THE BIOTECHNO LOGY IND USTRY

The Chakrabarty decision will add some pro-

tection for microbiological inventions by pro-

viding companies with an additional incentive

for* the commercial development of their inven-

tions, particularly in marginal cases, by lower-ing uncertainty and risk. A greater effect willresult from the new information disclosed inpatents on inventions that otherwise might have

been kep t secret indefinitely. Competitors andacademicians w ill gain new know ledge as wellas a new organism upon which to build. ThePatent office had deferred action on abou t 150

applications, while awaiting the Court’s deci-sion; as of December 1980, it was p rocessing ap -proximately 200 applications on micro-orga-nisms.

16*

Depending on the eventual num ber and im-portance of patented inventions that wouldhave otherwise been kept as trade secrets, theultimate effect of the d ecision on innovation in

the biotechnology industry could be significant.

16R[ +11 [ ) ‘l’[~~llll~v~l,, ,\ SSISlillll (:t)llllllissi[)ll(![’ 1’01” PillPlltS, [ 1.S. Pii l -

(1111 illl(l ‘ l ’ 1 . i i d o l l i i i ~ . k otl’i(x?, }](~t’sotliil (’otllllltllli($iit iou, J i l t ] . 8 , I !] & 1.

“1’11[’S(’ ill)pli(’iltiol)S i 1l(’lll(lt> ill)()(lt I ()() 011 ~[gllf]t i(’illl}r (’11-

gineerd m i (w tw s i l l l ( l iil)ollt 1 0 0 ” 011 (“llltlll’[~S ot” stl-;iil]s is(]lilt(~(l

1’1’0111 Ililt(ll’(’.

Conversely, if the Court had reached the op-posite decision, the industry would have beenheld back only moderately because of reason-ably effective alternative means of protection.

Impac ts o f the Court ' s dec is ion on the

p a t e n t l a w a n d t h e P a t en t

a n d T r a d e m a r k O f f i c e

The key rationale supporting the Court’sholding Chakrabarty’s m icrobe to be patentable

was th e fact that it was m anmade; its status as aliving organism w as irrelevant. The Patent Of-fice interprets this decision as also permittingpatents on micro-organisms found in nature butwhose useful properties depend on hum an in-tervention other than genetic engineering,

l7

e.g., if the isolation of a pure culture of amicrobial strain induces it to produce an an-tibiotic, that pure culture would be patentablesubject matter.

Because of the complexity, reproducibility,and mutability of living organisms, the decisionmay cause some p roblems for a body of law de-signed m ore for inanimate objects than for liv-ing organisms. It raises qu estions about theproper interpretation and application of the re-quirements for novelty, nonobviousness, andenablement. In addition, it raises questionsabout how broad the scope of patent coverageon imp ortant micro-organisms shou ld be and

about the continuing need for the two plant pro-tection Acts. These uncertainties could result inincreased litigation, making it more difficult andcostly for owners of patents on living organism!to enforce their rights.

The complexity of living m atter w ill make idifficult for anyone examining the invention todeterm ine if it meets the requiremen ts for novelty, nonobv iousness, and enablement. Micro-organisms can have different characteristics indifferent environments.’ Moreover, microbictaxonomists often differ on the precise classifi-

cation of microbial strains. Even a fter expensivetests, uncertainty may still exist about whethera specific micro-organism is distinct from othterknown strains; scientists do not have complete

‘‘1 hid, J i~ l l . 7, 19 S 1.




I s s u e a n d O p t i o n s

I SSU E : T o w h a t ex t en t co u ld Co n g r es sp r o v id e f o r o r p r o h ib i t t h e p a t -

en ta b i li t y o f l iv ing o rgan i sm s?

In its Chakrabarty opinion, the Supreme

Court stated that it was undertaking only thenarrow task of determining whether or notCongress, in enacting the patent statutes, hadintended a manmade micro-organism to be ex-clud ed from paten tability solely because it wasalive. Moreover, the opinion specifically invitedCongress to overrule the decision if it disagreedwith the Cou rt’s interpretation.

Congress has several options. It can act to re-solve the questions left unanswered by theCourt, overru le the decision, or develop a com-prehensive statutory approach. Most important-

ly, Congress can draw lines; it can decide whichorganisms, if any, should be patentable.

OPTIONS

A: Congress could maintain the status quo.

Congress could choose not to address theissue of patentability and allow the law to bedeveloped by the courts. The advantage of thisoption is that issues will be addressed as theyarise in the context of a tangible, nonhypo-thetical case. Some of the issues raised in thedebate on patenting may turn out to be irrel-

evant as the technology and the law develop.Moreover, many of the uncertainties raised byth e Chakrabarty decision regarding provisionsof the patent law other than section 101 may beincapable of statutory resolution. The complexi-ty of living organisms an d the increase in kn owl-edge of molecular genetics will raise such broadand varied questions that legal interpretationsof whether a particular biological inventionmeets the requirements of novelty, nonobvious-ness, and enablement will best be done on acase-by-case basis by the Patent Office and theFederal courts.

There are two disadvantages to this option.First, a uniform body of law may take time todevelop, since judicial decisions about new legalquestions by different Federal courts may ini-

tially conflict. Second, the Federal judiciary isnot designed to take sufficient account of thebroader political and social interests involved.

B: Congress could pass legislation dealing with

the specific legal issues raised by the Court's decision.

Many of the legal questions do not readilylend themselves to statutory resolution. How-ever, three questions are fairly narrow andwell-defined and may therefore be better re-solved by statute: 1) Is there a continuing needfor the plant protection Acts if plants can bepatented u nd er section 101? 2) If there is a con-tinuing need for these Acts, could they be ad-ministered better by one agency? 3) Shou ld th edefinition of infringement be clarified by

amending section 271 of the Federal PatentStatu tes (title 35 U. S. C.) to includ e rep roductionof a patented organism for the purpose of sell-ing it?

Congressional action to clarify these issueswould provide direction for industry and thePatent Office, and it would obviate the need fora resolution through costly, time-consuming lit-igation. Lessening the chances of litigation orthe chances of a patent being declared invalidwill provide some stimu lation for innovation bylessening the risks in commercial development.In addition, Congress could determine that theplant protection Acts could be better admin-istered by one agency or should be incorporatedunder the more general provisions of the patentlaw; if so, some adm inistrative expenses p rob-ably could be saved.

C: Congress could mandate a study of the plant protection Acts.

Two statutes, the Plant Patent Act of 1930and the Plant Variety Protection Act of 1970,grant ownership rights to plant breeders whodevelop new and distinct varieties of plants.

They could serve as a model for studying thebroader, long-term potential imp acts of paten t-ing living organ isms. An empirical stud y of theimpacts of the plant protection laws has notbeen done. Such a study would be timely, not



Ch. 12—Patenting Living Organisms . 253

only because of the Chakrabarty decision, butalso because of allegations that the Acts have en-couraged the p lanting of un iform v arieties, lossof germplasm resources, and increased concen-tration in the plant breeding industry. In addi-tion, information about the Acts’ affect on in-

novation and competition in the breeding in-dustry would be relevant to this aspect of thebiotechnology industry. However, it may be ex-tremely difficult to isolate the effects of theselaws from the effects of other factors.

D: Congress could prohibit patents on any living organism or on organisms other than those already subject to the plant protection Acts.

By prohibiting patents on any living orga-

nisms, C o n g r e s s w o u l d b e a c c e p t i n g t h e

arguments of those who consider ownership

rights in living organisms to be immoral, or whoare concerned about other potentially adverseimpacts of such patents. Some of the claimed

impacts are: 1) patents would stimulate the de-

velopment of molecular genetic techniques,

which will eventually lead to h uman genetic en-

gineering; 2) patents contribute to an atmos-

phere of increasing interest in commercializa-tion, which will discourage the open exchange

of information crucial to scientific research; and

3) plant patents and protection certificates have

encouraged the planting of uniform varieties,loss of germplasm resources, and increasing

concentration in the plant breeding industry.Also, by repealing the plant Acts, Congress

would be reversing the policy determination it

made in 1930 and in 1970 that ownership rightsin novel variet ies of plants would st imulate

plant breeding and agricultural innovation.

A prohibitory statute would have to deal with

those organisms at the edge of life, such asviruses. Although there are uncertainties and

disagreements in classifying some enti t ies asliving or nonliving, Congress could be arbitraryin its inclusions and exclusions, so long as it

clearly dealt w ith all of th e difficult cases.This statute by itself would slow but not stop

th e<development of molecular genetic tech-

niques and the biotechnology industry because

there are several good alternatives for maintain-

ing exclusive control of biological inventions:

maintaining organisms as trade secrets; patent-

ing microbiological processes and their prod-

ucts; and patenting the inanimate components

of a genetically engineered micro-organism,

such as plasmids, which are the crucial ele-

ments of the technique anyway. The develop-

ment would be slowed primarily because infor-mation that might otherwise become public

would be kept as trade secrets. A major conse-

quence would be that desirable products wouldtake longer to reach the market. Also, certainorganisms or products that might be marginallyprofitable yet beneficial to society, such as somevaccines, would be less likely to be d eveloped.In such cases, the recovery of developmentcosts would be less likely without a patent toassure exclusive marketing rights.

Alternatively, Congress could overrule the

Chakrabarty decision by amending the patentlaw to prohibit patents on organisms other than

the plants covered by the two statutes men-

tioned in option C. This would demonstratecongressional intent that living organisms could

be patented only by specific statute and alleviate

concerns of those wh o fear the “slippery slope. ”

E: Congress could pass a comprehensive law covering any or all organisms (except humans).

This option recognizes the fact that Congresscan draw lines where it sees fit in this area. It

could specifically limit patenting to micro-orga-nisms or encourage the breeding of agricul-

tura l ly impor tant animals by grant ing patent

rights to breeders of new and distinct breeds.

Any fears that such patents would eventuallylead to patents on hum an beings would be un -found ed, since the 13th am endment to the con-stitution, which abolished slavery, prohibitsownership of hu man life.

The statute would have to define included orexcluded species with precision. Although thereare taxonomic uncertainties in classifying or-

ganisms, Congress could arbitrarily include orexclude borderline cases.

A statute that permitted patents on several

types of organisms could be modeled after the

Plant Variety Protection Act—e.g., it should

cover organisms that are novel, dist inct , and




uniform in reproduction; such terms wouldhave to be d efined, Infringemen t should includ ethe unauthorized reproduction of the orga-nism-although reproduction for researchshould be excluded to allow the development of new varieties. In fact, consideration should be

given to covering in one statute plants and allother organisms that Congress desires to be pat-entable. This would provide the advantage of comprehensiveness and uniform treatment; itcou ld a l so address the p rob lem s d i scussed

und er option B.

The impact of this law cannot be assessed

precisely. A comprehensive statute would stim-

ulate the development of new organisms and

t h ei r p ro d u ct s a n d w ou l d en c ou r a ge d i s -

semination of technical information; however,

s u c h a s t a t u t e i s n o t e s s e n t i a l t o t h e d e -

velopment of the biotechnology industry, sinceincentives and alternative means for protection

already exist. The secondary impacts on society

of the legislat ion are even harder to assess

because of the scarcity of data from which todraw conclusions. The policy judgments will

have to be mad e by Congress after it weighs the

opinions o f t h e v a r i o u s in t e res t g roups .

Through legislation, Congress has the chance to

balance competing views on this controversial

issue and, if necessary, to alleviate the primary

concerns about the long-term impacts of the

decision–that higher organisms will inevitably

be patented.



C h a p t e r 1 3

G e n e t i c s a n d S o c i e t y



C h a p t e r 1 3

PageGenetics and Modern Science . . . . ...........257 The “Public” and “Public Participation” . . . . . . . .261Special Problems Posed by Genetics . .........258 Issues and Options . . . . . . , . . . . . . . . . . . . . . . . 261

science and Society. . . . . . . . . . . . . . . . . . . . .. .259 Bibliography: Suggested Further Reading , . . . . .26.5



Ch ap te r 1 3

G e n e t i c s a n d S o c i e t y

G e n e t i c s a n d m o d e r n s c i e n c e

In 1979, the organization for Economic Coop-

eration and Development (OECD)* published a

survey of mechanisms for settling issues involv-.ing science and technology in its member coun-

tries. I The OECD report noted that:2

Science and technology . . . have a n um ber of distinguishing characteristics which cause spe-cial problems or complications. One is ubiquity:they are everywhere. They are at the forefrontof social chan ge. They not only serve as agents

of change, but provide the tools for analyzing

social change. They pose, therefore, special 3.

challenges to any society seeking to shape itsown future and not just to react to change or to

the sometimes undesired effects of change.

After surveying member countries, OECD

identified six factors that d istingu ish issues inscience and technology from other p ublic con-troversies.

1.

2.

The rapidity of change in science and technology often leads to concern. The scienceof genetics is one of the most rapidly ex-

panding areas of human knowledge in the

world todayv. And the technology of geneticsis causing quick and fundamental changes

on a variety of fronts. The news media

have consistently reported developments

in genetics, often with front-page stories.Consequently, the public has become in- 4.creasingly aware of developments in genet-ics and genetic technologies and the speedwith w hich knowledge in the field is gath-ered and applied. Many issues in today's science and technology are entirely new. Protoplasm fusion, re-

“’1’he m e m b e r s of’ OE(;D are: Auslra]ia, Austria, Belgium,

(;ilni]da, l)enmark, Finland, France, West Germany, Greece, lce-Iand, l r ~ l i i n d , I ta l y , J a p a n , I , u x em hou r g , the Netherlands, New

Zeiiliind, Norway, Porttl~i\l, Spain, Sweden, Switzerland, Turkey,

[he (fnited Kingdom, iitld th e LJnited States.1( ;uild K. Nichols, ‘1’echnologv on Trial: Public Participation in De-

cision-Makin~ Related to Science and Technology [Paris: organiza-tion fo r kkwnomic (kmpera t ion and Development, 1979).

‘lt~id., p. 16.

combinant DNA (rDNA), gene synthesis,chimeras, fertilization of mammalian em-bryos in vitro, and the successful introd uc-tion of foreign genes into mammals werethe subjects of science fiction until a fewyears ago. Now they appear in new spapersand popular magazines. Yet the genera]public’s understanding of these phenom-ena is limited. It is difficult for people toevaluate competing claims about the dan-gers and benefits of this new technology.The scale, complexity, and interdependence

among the technologies are greater thanpeople suspect. As in other fields, applica-tions of biological technology often dependon parallel developments in areas that pro-vide critical sup port systems. Breakdow nsin these system s are often as limiting as fail-ures in th e new technology itself. In otherparts of this report for example, sophis-ticated breeding systems in farm animalsand large-scale fermentation processes forsingle-cell cultures are described. Besidesthe biological technology requ ired to su p-port these systems, precise computerized

operations are required to ensure purity,safety, and process control in fermentationand to provide the population statisticsnecessary for breeding decisions.Some scientific and technological achieve-ments may be irreversible in their effects.

Because living organisms reproduce, somefear that it will be impossible to containand control a genetically altered organismthat find s its way into the environment an dproduces undesirable effects. Scenarios of escaping organisms, pand emics, and care-less researchers are often d raw n by critics

of today’s genetics research. The intention-al release of recombinant organisms intothe environment is a related issue that willneed to be resolved in the future.

Another example of irreversibility,brought about by the demands placed on

257



Ch. 13—Genetics and Society . 259

entist or physician for daring to “play God?” IS it

because we have forgotten the Semitic (biblical)

conception of creation as God’s ongoing col-

laboration with man? Creation is our God-given

role, and our task is the ongoing creation of the

yet unfinished, still evolving nature of man.

Man has played God in the past, creating awhole new artificial world for his comfort and

enjoyment. Obviously we have not always dis-played the necessary wisdom and foresight in

that creation; so it seems to me a waste of time

and energy for scientists, ethicists, and laymen

alike to beat their breasts today, continual ly

pleading the ques t ion of whether or not we

have the wisdom to play God with human na-

ture and our future. It is obvious we do not, and

never will, have all the foresight and prudence

we need for our task. But I am also convinced

that a good deal of the wisdom we lack could

have been in our hands if we had taken serious-

ly our human vocat ion as t ranscendent crea-

S c i e n c e a n d so c i e t y

The public’s increasing concern about the ef-

fects of science and technology has led to de-

mands for greater participation in decisions on

scientific and technological issues, not only in

the Uni ted S ta tes but throughout the wor ld .

The demands imply new challenges to systemsof representative government; in every West-

ern country, new mechanisms have been de-vised for increasing citizen participation. An in-

creasingly informed population, skilled at exert-ing influence over policymakers, seems to be a

strong trend for the future. The media has

played an important role in this development,

reporting both on breakthroughs in science and

technology and on accidents, pollution, and the

side-effects of som e technologies.

one result has been the growing politiciza-

tion of science and technology. While perhaps

misunderstanding the nature of science as a

process, the public has become disenchanted byrecent accidents associated with technology, by

experts who openly disagree with one another,

and by the selective use of information by some

scientific supporters to obtain a political objec-

tive. The public has seen that technology affects

tures , c rea tures or iented toward the future

(here and hereafter), a future in which we arecocreators.

Genetics thus poses social di lemmas that most

other t echnologies based in the physical sc i -

ences do not. Issues such as sex selection, the

abortion of a genetically defective fetus, and invitro fert i l ization raise conflicts between in-

dividual rights and social responsibility, and

they challenge the religious or moral beliefs of

many. Furthermore, people sense that genetics

will pose even more difficult dilemmas in the fu-

ture. Although many cannot fully articulate the

basis for their concern, considerations such asthose discussed in this section are cited. The

strong emotions aroused by genetics and by the

questions of how much and what kind of re-

search should be done are at least partly rooted

in deeply held human values.

the distribution of benefits in society; it canhave unequal impacts, and those who pay orwh o are most in need are not necessarily always

those who benefit.

A national opinion survey of a rand om sam-ple of 1,679 U.S. adults conducted for the Na-tional Commission for the Protection of HumanSub jects of Biomed ical and Behav ioral Research

4

made clear that there is public doubt concern-ing equity. Sixty percent of those polled felt that

new tests and treatments deriving from medical

research are not equally accessible to all Amer-

icans. Seventy percent felt that those most likely

to benefit from a new test or treatment of lim-

ited availability were those who could pay for itor wh o knew an important d octor. This shouldbe compared with the 85 percent w ho felt that anew test or treatment should be available tothose who ap ply first or who a re most in need.

4’4S~)[W’iill SIU(IJ’, IIllpli(’iitioll!i 01” ,ll(lt’illl(’t?$i iil f}ioll)fl(li(’ill illl[l B(’-

l~il\ iol’ill Ros ( ’ i i l ’ ( . l l , ’” R( ’ l )o l> t i i l l [ l R( ’ ( ” o t l ] t l l ( ’ t l ( l i i l i o l l s ” 01 ” th (~ Ni i t io l l i i l

( ‘(mltllissioll t“ol” Illf> l}lx)t(~(liotl ot” Illllllilll Slll)jo(’ls ot” lliolll(~(li(’ill

ilt~(l l\(*ilit\’iol. ii I K(’~f~iit.(’11, I III I“;\\’ I)(ll)li(’iil ion N’(). (OS) 78-()() 15.




T h e “ p u b l i c ’)a n d ‘(p u b l ic p a r t i c ip a t i o n ”

These are terms with vastly different mean-ings to different people. Some take “the public”to mean an organized public interest group;

others consider su ch group s the “professional”public and feel they have agendas that differfrom those of the less organized “general” pub-lic. As OECD stated :

8

Public participation is a concept in search of a

definition. Because it means different things to

different people, agreement on what constitutes

“the public” and what delineates “participation”

is difficult to achieve. The pu blic is not of course

homogeneous; it is comprised of many hetero-

geneous e lements , inte res ts , and p reoccupa-

tions. The emergence over the last several dec-

ades of new and sometime vocal special interest

groups, each with i ts own set of competingclaims and demands, attests to the inherent dif-

ficulty of achieving social and political consen-

sus on policy goals and programmed purporting

to serve the common interest.

‘ N k > h o l s , op . ( ’ i t . , p . 7.

Because publics differ with each issue, no def-inition will be attemp ted h ere. It is assumed that“the pu blic” is demand ing a greater role in de-

cisions about science and technology, and that itwill continue to do so. The different publics thatcoalesce around different issues vary w idely in

their basic interests, their skills, and theirultimate objectives. They are the groups thatwill be heard in the widening debate aboutscientific and technological issues, and are partof what has been called the “social system of science. ”

9

The public has already become involved inthe decisionmaking process involving geneticresearch. As the science develops, new issues inwhich the public will demand involvement willarise. The question is therefore: What is thebest way to involve the public in decision-making?

“J. Al. Zinl~ln, fublic K/u)w/@e (( ~iitlll)l’idgt~: (~ilt)~l)l.i(l~(l [ ll]i\wr-

sil.v PIWS, 1 !)68).

I s su e s a n d O p t io n s

Three issues are considered. The first is anissue of process, concerning public involvement

in policymaking; the second is a technical issue;and the third reflects the complexity of someissues associated with genetics that may arise inthe future.

ISSUE: H o w s h o u ld t h e p u b l i c b e i n -v o l v e d i n d e t e r m i n i n g p o l i c y r e -l a t ed t o n ew ap p l i ca t i o n s o f ge -n e t i c s ?

The question as to whether the public shouldbe involved is no longer an issue. Groups de-mand to be involved when people feel that their

interests are threatened in ways that cannot beresolved by representative democracy,

The more relevant questions are whethercurrent mechanisms are adequate to m eet pub-lic desires to participate and whether a de-

liberate effort should be m ade to increase pub-lic knowledge. The last can only be accom-

plished by educating the public and increasingits exposure both to the issues and to how peo-ple may be affected by different decisions.

OP TIONS :

A.

B.

Congress could specify that the opinion of the public must be sought in formulating all major policies concerning new applications of genetics, including decisions on funding of specific research projects. A “public participation statement” could be mandated for all such decisions.

Congress could maintain the status quo, al low-

ing the public to participate only when it decides to do so on its own initiative.

If option A w ere followed, there w ould be n ocause for claiming that public involvement was

76-565 0 - 81 - 18




inadequate (as occurred after the first set of Guidelines for Recombinant DNA Researchwere prom ulgated). However, option A can beimplemented in two ways. In the first, the op-portunity for public involvement is always pro-vided, bu t need not be taken if there is no public

interest in the topic. In the second, public in-volvement is required. A requ irement for pu blicinvolvement would pose the problem that if thepublic does not wish to participate in a par-ticular decision, then opinion will sometimes besought from an uninterested (and thereforeprobably uninformed) pu blic simp ly to meet therequirement. Option A poses additional prob-lems: What is a “m ajor” policy? At wh at stagewould public involvement be required—onlywhen technological development and applica-tion are imminent or at the stage of basicresearch? Finally, it should be noted th at un der

option A, if the public’s contribu tion significant-ly influences policy, the trend away from deci-sionmaking by elected representatives (rep-resentative democracy) and toward decision-making by the people directly (“participatory”dem ocracy) may be accelerated.

Option B would be less cumbersome andwou ld perm it the establishment of ad hoc mech-anisms when necessary. On the other hand, bythe time some issues are raised, strong vestedinterests would already be in place. The grow-ing role of single-issue adv ocates in U.S. politics,

and their skill in influencing citizens and policy-makers, m ight abor t certain scientific develop-ments in the future.

Regardless of which option is selected, itwould be desirable to encourage differentforms of structuring public participation and toevaluate the su ccess of each method . Many d if-ferent approaches to public participation havebeen tried in the United States and WesternEurope in attempts to resolve conflicts overscience and technology. Some h ave w orked bet-ter than others, but most have had ratherlimited success.

l0Because public demands for

involvement are not likely to d iminish, the best

1(J[ )i)l.[lll~v N[?l~ill iII)d h~ i[:hiie! Pollil[!k, “}’1’01)1(?111s iIINi 1%’(){:(?-

dUI’(W in I}W 11(’~iiiiitioll 01 ‘ l ’ t ’ ( s l l l lo lo~ i ( s i l l ” Risk ,“ in SOCield Risk As-

S~S.WTI~~I, Il. Schwing, illld W . ,III)(YIvJ ((+(1s.1 (N(?w Yot’k: Pl(!I~LII]]

1%’ss, 198(1).

ways to accommodate them need to be iden-tified.

I SSU E : H o w can th e l ev e l o f p u b l i ck n o w le d g e c o n c e r n i n g g e n e t i c s

a n d i t s p o t e n t i a l b e r a i s e d ?

If public involvement is expected, an in-formed public is clearly desirable. Increasingthe treatment of the subject, both within andoutside the traditional educational system, is theonly way to accomplish this.

Within the trad itional edu cational system, atleast some educators feel that too little time isspent on genetics. Some, such as members of the Biological Sciences Curriculum Study Pro-gram, are considering increasing the share of the curriculum devoted to genetics. Becausescience and technology cause broad changes in

society, not only is a clearer perception of genetics in particular needed, but more under-standing of science in general. For about half the U.S. population, high school biology is theirlast science course. Educators must focus onthis course to increase public understanding of science. Because students generally find peoplemore interesting than rats, and because hu mangenetics is a very popular topic in the highschool biology course, educators responsible forthe Biological Sciences Curriculum Study Pro-gram are considering increasing time spent onits study in hopes of increasing public knowl-

edge not only of genetics but of science in gen-eral.

At the university level, more funds could beprovided to develop courses on the relation-ships between science, technology, and society,wh ich could be designed both for students andfor the general public.

Several sources outside the traditional schoolsystem already work to increase public under-standing of science and the relationships be-tween science and society. Among them are:

q

Three programs developed by the NationalScience Foundation to improve publicunderstanding of and involvement in sci-ence: Science for the Citizen; Public Under-stand ing of Science; and Ethics and Valuesin Science and Technology.



Ch. 13—Genetics and Society q 263

q

q

q

q

Science Centers and similar projects spe-cifically designed to present science infor-

mation in an app ealing fashion.

New magazines that offer science informa-

tion to the lay reader-another indication

of increasing interest in science.

Television programs dealing with scienceand technology. Examples are the two PBSseries, NOVA an d Cosmos, and the BBC

series, Connections. CBS has also begun anew series called The Universe.Television programs dealing with social

issues and value conflicts. Particularly in-

teresting is the concept behind The Baxters.[n this half-hour prime time show, the net-

work provides the first half of the show,

which is a dramatization of a family in con-

flict over a social or ethical issue. The sec-

ond half of the show consists either of a dis-

cussion about what has been seen or of comments from people who call in.

One interesting possibility would be to com-

bine a series of Baxter-type episodes on genetic

issues with audience reaction using the QUBE

system, a two-way cable television system in

Columbus, Ohio (now expanding to other cities).

In this system, television viewers are provided

w i th a s im ple dev ice tha t enab les them to

answer questions asked over the television. A

computer tabulates the responses, which caneither be used by the s tudio or immediate ly

transmitted back to the audience. QUBE permitsits viewers to do comparison shopping in dis-

count stores, take college courses at home, and

provide opinion to elected officials. It could be

effectively combined with a program like The

Baxters, to study social issues. If several such

programs on genetics were shown to QUBE sub-

scribers, audience learning and interest could

be measured.

Any efforts to increase public understanding

should, of course, be combined with carefully

designed evaluation studies so that the effec-

tiveness of the program can be assessed.

OP TIONS :

A. Programs could be developed to increase public understanding of science and the rela -

tionships between science, technology, and society.

Public understanding of science in today’s

world is essential, and there is concern about

the adeq uacy of the pub lic’s kn owledge.

B. Programs could be established to monitor thelevel of public understanding of genetics and of science in general and to determine whether

public concern with decisionmaking in science

and technology is increasing.

Selecting this option would indicate thatthere is need for additional information, andthat Congress is interested in involving the p ub-lic in d eveloping science p olicy.

C. The copyright laws could be amended to permit schools to videotape television programs for educational purposes.

Under current copyright law, videotaping

television programs as they are being broadcast

may infringe the rights of the program’s owner,

generally its producer. The legal status of such

tapes is presently the subject of litigation. As a

matter of policy, the Public Broadcasting Serv-ice negotiates, with the producers of the pro-

grams that i t broadcasts, a l imited right forschools to tape the program for educational

uses. This permits a school to keep the tape for agiven period of time, most often one week, after

which it must be erased. otherwise, a school

must rent or purchase a copy of the videotape

from the owner.

In favor of this option, it should be n oted that

many of the programs are made at least in part

with public funds. Removing the copyright con-straint on schools would make these programsmore available for another pu blic good, edu ca-tion. On the other hand , this option could h avesignificant economic consequences to the copy-

right owner, whose market is often limited toeducation] institutions. An ad hoc committee of

producers, educators, broadcasters, and talent

unions is attempting to develop guidelines in

this area.




ISSUE: Should Congress beg in p repar -in g n o w to r e s o lv e i s s u es t h a th a v e n o t y e t a r o u s e d m u c h p u b -l i c d e b a t e b u t t h a t m a y i n t h ef u t u r e ?

As scientific understanding of genetics and

the ability to m anipu late inherited characteris-tics develop, society may face some difficultquestions that could involve trad eoffs betweenindividu al freedom and societal need. This willbe increasingly th e case as genetic technologiesare applied to humans. Developments are oc-curring rap idly. Recombinan t DNA technologywas developed in the 1970’s. In the spring of 1980, the first application of gene replacementtherapy in mammals succeeded. Resistance tothe toxic effect of methotrexate, a dru g u sed incancer chemotherapy, was transferred to sen-sitive mice by substituting the gene for resist-

ance for the sensitive gene in tissue-culturedbone marrow cells obtained from the sensitivemice. Transplanted back into the sensitive mice,the bone marrow cells now conferred resist-ance to the drug.

11In the fall of 1980, the first

gene substitution in humans was attempted.l2

Although this study was restricted to non-human applications, many people assume fromthe above and other examples that wh at can bedone w ith lower animals can be done w ith hu-mans, and will be. Therefore, some action mightbe taken to better prep are society for decisions

on the application of genetic technologies tohumans.

OP TIONS :

A. A commission could be established to identify central issues, the probable time-frame for application of various genetic technologies to humans, and the probable effects on society, and to suggest courses of action. The commission might also consider the related area of how participatory democracy might be combined with representative democracy in deci-

sionmaking.

B. The life o f the President's Commission-for theStudy of” Ethical Problems in Medicine and Biomedical and Behavioral Research could beextended for the purpose of addressing theseissues.

The n-member Commission was established

by Public Law 95-622 in November 1978 andterminates on December 31, 1982. Its purpose isto consider ethical and legal issues associatedwith the protection of human subjects in re-search; the definition of death; and voluntarytesting, counseling, information, and edu cationprograms for genetic diseases as well as anyother appropriate topics related to medicineand to biomedical or behavioral research.

In July and September 1980, the Commissionconsidered how to respond to a statement fromthe general secretaries of the N ational Council

of Churches, the Synagogue Council of America,and the United States Catholic Conference th atthe Federal Government sh ould consider ethicalissues raised by genetic engineering. The re-quest was prom pted by the Supreme Court d eci-sion allowing patents on “new life forms. ” Thegeneral secretaries stated that “no governmentagency or committee is currently exercisingadequate oversight or control, nor addressingthe fundamental ethical questions (of geneticengineering) in a major way, ” and asked that thePresident “provide a way for representatives of a broad spectrum of our society to consider

these matters and advise the government on itsnecessary role, ”

13

After testimony from various experts, theCommission found that the Government is al-ready exercising adequ ate oversight of the “bio-hazard s” associated with rDNA research and in-du strial prod uction. The Commission d ecided toprepare a report identifying what are and arenot realistic problems. It will concentrate on theethical and social aspects of genet ic technologythat are most relevant to medicine and b io -

medical research.

The Commission could be asked to study theareas it identifies and to broaden its coverage to

1 ‘J(? i i l l [,. h l i l ] s ~ , “ ( k i l t ? ‘ l ’ l s i t l ] s f (? l ’ ( ;h ’en ii New Twi s t , ” S c i e n c e

~()~:~~, ,\pril 1980, p. 386.

‘2(;i!]il l \ i ii ’ i Kolii[ii illl[{ Ni($holiis Wit(It?, “ t{tlt]liiil (kIM ‘ l ’ ls~iill]l~t~t

Stirs N(IW I)(II);I[(I, ” Science 2 10:24, octolwr 1980, p. 407.

t~slillell)ellt I)v the gellel.il] s~t:l’~t~l.i~s, (1 .S. ‘C:alholic [k)ll~t?l’-

(vM:[?, or”igi[ls, N( I l)()(!tltllelltiilm~ Service, vol. l o , N(.). 7 , JLII.V Li,

1 !)/J().




addit ional areas. This would require that i ts

term be extended and that additional funds be

appropriated. The Commission operated on $1.2million for 9 months of fiscal year 1980 and $1.5million for fiscal year 1981. Given the complexi-ty of the issues involved, the adequacy of this

level of fund ing should be reviewed if add itionaltasks are undertaken.

A potential disadvantage of u sing the existing

Commission to address societal issues associated

with genetic engineering is that a number of

issues already exist and more are likely to ap-pear in the years ahead. Yet there are also other

issues in medicine and biomedical and be-havioral research not associated with geneticengineering that need review. Whether all

these issues can be addressed by one Commis-sion should be considered. There are obviousadvantages and disadvantages to two Comm is-

sions, one for genetic engineering and one forother issues associated with medicine and bio-medical and behavioral research. Commentsfrom the existing Commission would assist inreaching a decision on the most appropriatecourse of action.

B i b l i o g r a p h y : s u g g e s t e d f u r t h e r r e a d i n g

Dobzhansky, Theodosium, Genetic Diversity and Hu- man Equality (New York : Basic Tool s, 1973).

A discussion of conflicts between the findings

of science and democratic social goals. Detailed

coverage of the scientific basis for present de-

bates about intelligence and the misconceptions

often involved in genetic v. environmental deter-

minants of certain human traits.

Francoeur, Robert T., “We Can - We Must: Reflec-

tions on the Technological lmperative, ” Theobgi-

cal studies 33 (#3): 428-439, 1972,

Argues that man is a creator by virtue of hisspecial position in nature, and that humans must

participate in deciding the course of their evolu-tion.

Goodfield, June, Plaving God: Genetic Engineering and

the Manipulation of Life (New York : Ha rpe r Col -

ophon Books, 1977).

Discusses the benefits, problems and potential

of gene t ic engineer ing. Descr ibes the mora l

dilemmas posed by the new technology. Suggests

that the “social contract” between science and

society is being “renegotiated.

Harmon, Willis, An Incomplete Guide to the Future

(New York: Simon and Schuster, 1976).

Surveys how social attitudes and values havechanged throughout history and how they may

be changing today. Argues that mankind is in the

midst of a transition to new values that will affect

our world view as profoundly as did the industri-

al revolution in the 19th century.

Holton, Gerald, and William A. Blanpeid (eds.), Sci-ence and Its Public: The Changing Relationship(Boston: D. Reidel, 1976).

A collection of essays on the way science and

the society of which it is a part interact, and howthat interaction may be changing.

Hutton, Richard, Bio-Revolution: DNA and the Ethics

of Man-Made Life (New York: New American Li-

brary (Mentor), 1978).

Reviews the history of the debate about recom-binant DNA, discusses the scientific basis for the

new technologies, and discusses the changingrelationship between science and society. Sug-

gests how the controversies might be resolved.

Monod, Jacques, Chance and Necessity (New York:

Alfred Knopf, 1971).

A philosophical essay on biology. Two seem-

ingly contradictor-v laws of science, the constan-

cy of inheritance (“necessity”) and spontaneous

mutat ion (“chance”) are compared with more

vitalistic and deontological views of the universe.

An affirmation of scientific knowledge as the on-ly “truth” available to man.

Nichols, K. Guild, Technology on Trial: Public Participa- tion in Decision-Making Related to Science and Technology (Paris: Organization for Economic Co-

operation and Development, 1971).

Reviews mechanisms that have been used bycountries in Europe and North America to settle

disputes involving science and technology.




Sinsheimer, Robert, “Two Lectures on RecombinantDNA Research,” in The Recombinant DNA Debate,D. Jackson and Stephen Stich (eds.) (EnglewoodCliffs, N. J.: Prentice Hall, 1979), pp. 85-99.

Argues for proceeding slowly and thoughtfullywith genetic engineering, for it potentially has

far-reaching consequences.

Tribe, Laurence, “Technology Assessment and the

Fourth Discontinuity: The Limits of Instrumental

Rat iona l i ty ,” Southern California Law Review 46

(#~): 617-660, June 1973.An essay on the fundamental task facing man-

kind in, the late 20th century: the problem of

choice of tools. New knowledge, especially from

biology, will increasingly offer options for tech-

nology, the use of which will cause changes in

human values.



A p p e n d i x e s

I-A. A Case Study of Acetaminophen Production . . . , . . . . . . . . . . . . . . . . . . . .269

I-B. A Timetable for the Commercial Production of Compounds Using

Genetically Engineered Micro-Organisms in Biotechnology . . . . . . . . . . . .275

I-C. Chemical and Biological Processes . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 292

I-D. The Impact of Genetics on Ethanol—A Case Study . . . . . . . . . . . , . . . . . .. 293

II-A. A Case Study of Wheat. , . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .304

II-B. Genetics and the Forest Products Industry Case Study. . . . . . . . , . . . . . . .307

H-C. Animal Fertilization Technologies. . . . . . . . . . . . . . . . . . . . . . . . , . . . . . . .309

III-A. History of the Recombinant DNA Debate . . . . . . . . . . . . . . . . . . . . .. ...315

HI-B. Constitutional Constraints on Regulation , . . . . . . . . . . . . . . . . . . , . . . . . .320

III-C. Information on International Guidelines for Recombinant DNA . . . . . , . . 322

IV. P l a nn i ng workshop Pa r t i c i pa n t s , O t he r Cont ra c t o r s a nd Cont r i bu t o r s , a nd

Acknowledgments. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . , , .329




Table l-A-5.-Labor Distribution for Production of APAP

Salary and wage cost

Category Man-hours per week Hourly rate $/week $/year

SupervisionGeneral manager . . . . . . . . . . . . . . . . . . . . . . . . .Superintendents. . . . . . . . . . . . . . . . . . . . . . . . . .Managers . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .Supervisors. . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Hourly rated employees, servicesLaboratory

Level I . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .Level II. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .Level Ill . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .Level lV . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Maintenance and engineeringLevel l . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .Level lI. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .Level lIl . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .Level IV . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Hourly rated employees, productionFermentation department

Level l . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .Level II . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Level Ill . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .Level lV . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Recovery departmentLevel l . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .Level II . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .Level Ill . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .Level IV . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Subtotal. ......, . . . . . . . . . . . . . . . . . . . . . . . . .

408080

320

8080

12040

240240240160

200240

8080

320400

80120

. . . . . . . .

20171512

10865

10865

108

65

10865

...,. . . . .

$ 8001,3601,2003,840

800640720200

2,4001,9201,440

800

2 , 0 0 01,920

480400

3,2003,200

480600

. . . . . . . . . . . . . . . . . . . . .Add overtime@ 6% x 1.5 . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Subtotal . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .Add fringe benefits @25%. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Total salaries and wages. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

$ 41,60070,72062,400

199,680

41,60033,28037.44010,400

124,80099,84074,88041,600

104,00099,840

24,96020,800

166,400166,40024,96031,200

$1,476,800132,912

$1,609,712402,428

$2,012,140

SOURCE:GenexCorp.

Table l-A-6.-Steam Requirements forProduction of APAP

Operation Lb/batch

Sterilization, fermenters, and seed tanks:Heating . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 52,100Holding . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 20,000

Sterilization, piping, and equipment (other) . . . 20,000Heating acetaminophen solution (recovery) ... 163,500Drying, turbo dryer. . ......................200,300General purpose usage . . . . . . . . . . . . . . . . . . . . 50,000

Total. . . . . . . . . . . . ......................505,900

Cost at $5.00/Mlb:Per fermenter batch =$ 2,530

Per year (100 batches) =$253,000

SOURCE:GenexCorp.



Appendix l-A—A Case Study of Acetaminophen Production . 273

Table l-A-7.–Electricity Requirements for Production of APAP

Units/batchConnected load HP kW (hours operation) kWh

Fermenters. . . . . . . . . . . . . . . . . . . . . . ........200Seed tanks . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 47.5Chillers . . . . . . . . . . . . . . . . . ................580Air compressor . . . . . . . . . . . . . . . . . . . . . . . . . .275H a r v e s t t a n k . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1 0 0Decanter centr i fuge. . . . . . . . . . . . . . . . . . . . . . .120Process tanks. . . . . . . . . . . . . . . . . . . . . . . . . . . .300Crysta l l iz ing tanks . . . . . . . . . . . . . . . . . . . . . . . .300Turbo dryer . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 30Cooling tower.. . . . . . . . . . . . . . . . . . . . . . . . . . . 40Pumps (est. =6@ 7.5) . . . . . . . . . . . . . . . . . . . . 45Lighting, instruments and general load. . . . . . . 25

Total kWh . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

@O.05/kWh = $3,469 per batch@100 batches/yr= $346,900 per year

14935

433205

90224224

22303419

14424

86115219

23144144144

21,456840

4,76317,630

8254,6804,2562,464

4,3204,8962,736

69,372

SOURCE:GenexCorp.

Table I-A.8.-WaterRequirements forProduction of APAP

Gal/batch

Fermentation . . . . . . . . . . . . . . . . . . . . . . . . . . 35,000Tower makeup . . . . . . . . . . . . . . . . . . . . . . . . . 63,000Process loss . . . . . . . . . . . . . . . . . . . . . . . . . . 100,000Chilled water makeup. . . . . . . . . . . . . . . . . . . 30,000Direct cooling . . . . . . . . . . . . . . . . . . . . . . . . . 50,000General use..... . . . . . . . . . . . . . . . . . . . . . . 25,000

Total . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 303,000 gal

Process Watergate =$1.00/M galscost = $303/batch

100 batches/yr =$30,300/yearSOURCE:GenexCorp.




Table l-A-9.—Equipment Costs forProduction of APAP

Receiving and batching area320,000 gal steel aniline storage tanks,

insulated and cooled - @ $47,000 . . . . . . . . . . . $ 341,000220,000 gal aluminum acetic acid storage

tanks, insulated and cooled - @ $71,300 . . . . . .

110,000 gal steel nitrogen storage tank withcontrols and instruments. . . . . . . . . . . . . . . . . .

110,000 gal steel lard oil storage tank,insulated and heated . . . . . . . . . . . . . . . . . . . . .

110,000 gal stainless steel Batch tank withprogrammable controller and agitator. . . . . . . . .

21,700 ft3 stainless steel Hopper bins withconveyors - @ $58,100 . . . . . . . . . . . . . . . . . . . .

1 Electric forklift truck . . . . . . . . . . . . . . . . . . . . . .

Fermentation and seed area1150 gal stainless steel seed vessel, fully

instrumented. . . . . . . . . . . . . . . . . . . . . . . . . . . .12,500 gal stainless steel seed vessel, fully

instrumented. . . . . . . . . . . . . . . . . . . . . . . . . . . .250,000 gal stainless steel fermenters, fully

instrumented with central control room -@ $399,000 . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Recovery area150,000 gal stainless steel process tank,

cooled, agitated and insulated . . . . . . . . . . . . .13,000 gal steel filter aid slurry tank

with agitator . . . . . . . . . . . . . . . . . . . . . . . . . . . .1 Stainless steel continuous decanter

centrifuge . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .2100,000 gal stainless steel process tanks,

insulated with external steam injection heater,pump and agitator- @ $333,000 . . . . . . . . . . . .

342,600

47,000

22,300

59,500

116,20011,400

125,000

169,000

798,000

195,000

11,300

167,000

666,000

120,000 gal stainless steel side-entering surgetank with agitator . . . . . . . . . . . . . . . . . . . . . . . . $ 56,100

350,000 gal stainless steel crystallizing tanks,insulated with heavy duty cooling coils andtop-mounted agitator - @ $195,000 . . . . . . . . . .

1 Stainless steel turbo tray dryer. . . . . . . . . . . . . .23,500 ft

3stainless steel hopper bins-

@ $66,000 . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .1 Bagging unit. . . . . . . . . . . . . . . . . . . . . . . . . . . . .4 Stainless steel finished product conveyors-

@ $12,000 . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .Auxiliary equipment31,500 c.f.m. reciprocating air compressors -

@ $168,000 . . . . . . . . . . . . . . . . . . . . . . . . . . . . .Laboratory and office equipment , . . . . . . . . . . . .Chillers, 500 ton total capacity . . . . . . . . . . . . . . .1 Cooling tower, 1,500 g.p.m.. . . . . . . . . . . . . . . . .35 Pumps and motors, various sizes. . . . . . . . . . .2 Dump trucks -$12,000 . . . . . . . . . . . . . . . . . . . . .Ventilation, general and spot - @ 7.5%

of equipment. . . . . . . . . . . . . . . . . . . . . . . . . . . .Piping, general, materials and installation -

@ 7.5% of equipment. . . . . . . . . . . . . . . . . . . . .Miscellaneous equipment (hand tools, etc.) -

@ 5°A of equipment . . . . . . . . . . . . . . . . . . . . . .

585,000

653,000

132,0002 0 , 0 0 0

4 8 , 0 0 0

4 9 8 , 0 0 06 5 0 , 0 0 05 7 5 , 0 0 02 1 0 , 0 0 0104 ,7002 4 , 0 0 0

5 8 3 , 7 9 1

5 8 3 , 7 9 0

3 8 9 , 1 9 4Total . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . $7,783,875

Annual charge for capital recovery over 10-yearperiod, with 12% interest compoundedannually ($7,783,875 x 0.17698) . . . . . . . . . . . . $1,377,590

SOURCE: Genex Corn.

Table I-A-10.–Buiiding Requirements for Production of APAPArea Gross space ft2/ft3 Unit valuea

cost

Central office . . . . . . . . . . . . . . . . . . . . 940 41.00b$ 38,540

Laboratories. . . . . . . . . . . . . . . . . . . . . 4,500 70.00b

315,000Warehouse . . . . . . . . . . . . . . . . . . . . . . 2,000/36,000 27.00b

54,000Batching . . . . . . . . . . . . . . . . . . . . . . . . 1,000/30,000 1.75 52,500Fermentation (including seed) . . . . . . 6,000/320,000 1.75 560,000Harvest, filter . . . . . . . . . . . . . . . . . . . . 3,500/169,000 1.75 295,750Processing, crystallization . . . . . . . . . 8,700/470,000 1.75 822,500Drying, finishing. . . . . . . . . . . . . . . . . . 5,000/270,000 1.75 472,500Warehouse, finished product. . . . . . . 11,000/200,000 27.00

b297,000

Auxiliary equipment. . . . . . . . . . . . . . . 4,300/154,000 1.75 269,500Maintenance, engineering. . . . . . . . . . 11,500/207,000 1.75 362,250

Total. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . $3,539,540

Amortization over 30 years@ 12% compound interest $439,399C

aunit values in cubic feet except where noted by “b.”bunit value in square feet.cAmortizatlon = 0.12414 X tOtal.

SOURCE: Genex Corp.



Append ix I -B

A T i m e t a b l e f o r t h e C o m m e r c i a l

P r o d u c t i o n o f C o m p o u n d s U s i n g

G e n e t ic a lly E n g in e e r e d Mic r o -

O r g a n i s m s i n B i o t e c h n o l o g y

Objec t ives

q The estimation of the prop ortions of various

g r ou p s o f co m m e r c ia l p r o d u c t s a n d p ro c e s s e s f o rwhich recombinant DNA (rDNA) technology could

be applicable.

q The construction of timetables to indicate plausi-ble sequenccs of commercial developments that

would result from the application of rDNA tech-nology.

A p p r o a c h e s

Th e following five industries were eva lua ted:

1. pharmaceutical,

2. agricultural,

3. food,

4. chemical, and5. energy .

The ma nufa c t u r i ng p roc e s se s t ha t woul d r e su l t

from the application of rDNA technology would be

based on fermentation technology, Therefore, a set

of parameters was developed to serve as a guide toassess the economics of applying fermentation tech-nology to the manufacture of products current ly

manufactured by other means.

The chemical industry generates a large number

of products that could be attributed to (and is in thisstudy) the other four industries cited, this particular

i ndus t ry wa s foc use d on more c l ose l y t ha n t he

others. The following factors were considered in

constructing the timetables showing the applicability

of rDNA technoloGY:q

q

q

q

q

th e cur ren t s ta te of the art of genetic engineer-

ing;

t h e c u r r e n t e c o n o m i c l i m i t a t i o n s o f f e r m e n t a -

t ion technology;the length of time to progress from a laboratoryprocess to the pilot plant to large-scale produc-

tion;

the plant construction time; a n d

t h e Government regulatory agency a p p r o v a l r e -

qu i re d (o f t he p roduc t s a nd ma nufa c t u r i ng

processes, not of the rDNA technology per se).

Sourc e s o f i n format ion

While much of the information compiled for thisreport was obtained from published sources, a con-siderable amount came from prior prop rietary stud-

ies performed by Genex Corp. In the latter case, in-formation is used that is not proprietary, although

the sources must remain confidential. In this connec-

tion Genex has had numerous discussions with the

technical and corporate management of more than

100 large companies (generally multibillion dollar

companies), concerning research interests, product

l ines, and market t rends. Production costs a r e ex-

t rapolated for four fermentation plants of various

sizes an d capabilities. (See table I-B-1.)

A group of Genex scientists, consisting of a bio-

chemical engineer, two organic chemists, a biochem-

ist, and four molecular geneticists rated the feasibili-

ty of devising micro-organisms to produce various

chemicals in accordance with the fermentation con-ditions sp ecified in table I-B-1. For tho se chem icls

that appeared to be capable of being produced mi-

crobiologically, dates were assigned f o r t h e t i m e s

when the necessary technology would be achieved in

the laboratory. By combining both technical and eco-

nomic factors, it then became possible to project a

timetable for commercial production. (See table

I-B-2.)

It should be emphasized that an extremely con-

servative approach was taken in considering fermen-

tation economics over the next 10 years. only the rel-

a t ive ly poor economics of convent iona l batch fer-

mentation was c o n s i d e r e d . i m m o b i l i z e d c e l l p r o c -

esses were projected to be 15 years away, and eventhen, the incremental cost savings projected (see

table I-B-1) are lower than the incremental cost sav-

ings currently obtained with immobilized cell proc-

esses. The assumptions made here, however, did in-

clude reasonably high product yields and highly effi-

275




Table l- B-5.-Food Products Table l- B-6.-Agricultural Products

Product category End use

Amino acidsGlutamate . . . . . . . . . . . .

Cysteine . . . . . . . . . . . . .

Aspartame. . . . . . . . . . . . .VitaminsVitamin C. . . . . . . . .

Vitamin D. . . . . . . . .

AromaticsBenzoic acid . . . . . .

AliphaticsPropionic acid. . . . .

Short peptidesAspartame . . . . . . . .

EnzymesRenninAmyloglucosidase.a-amylase . . . . . . . .

Glucose isomerase.

Nucleotides5

1-IMP

a. . . . . . . . . . .

51-GMP b . . . . . . . . . .

,.. .

. . . .

. . . .

,.. .

,.. .

. . . .

. . . .

. . . .

. . . .

Food enrichment agent,flavoring agent

Food enrichment agent,manufacturing processes

Flavoring agent

Food additive, foodenrichment agent

Food enrichment agent

Food preservative

Food preservative

Artificial sweetener

Manufacturing processesManufacturing processesFood enrichment agent,

manufacturing processesManufacturing processes:sweetener

Flavoring agent. . . . Flavoring agent

%’-lnosinic acid.b51-guanylic acid.

SOURCE: Genex Corp.

al l the R&D start ing from the current knowledge

base necessary to demonstrate that the desired com-

pounds can be biological ly produced fi rst in the

laboratory and then in the pilot plant at commercial-

ly desirable yields and reaction efficiencies. The

timetable does not consider delays caused by con-struct ion of new faci l i t ies nor delays required to

ob t a i n Gove rnme nt r e gu l a t o ry a pprova l o f ne w

products.

It should be noted that in the technical data charts,

when glucose is listed as an alternate precursor by

fermentation, other carbohydrates, e.g., cellulose

and cornstarch, could be used. Moreover, if glucosewere the precursor of choice, the actual feedstock

would probably be a commodity like molasses as op-

posed to pure glucose.

S u m m a r y

Over 100 compounds represent ing 17 differentproduct categories that span the five industries

under evaluation are represented in table I-B-10. Th ecurrent mark et value of all these prod ucts exceeds$27 billion. One particular compound, methane, ac-

counts for over $12 bi l l ion. The e ve n-numbe re d


Amino acidsLysine. . . . . . . . . . . . . . . . . . . . .Methionine. . . . . . . . . . . . . . . . .Threonine. . . . . . . . . . . . . . . . . .Tryptophan. . . . . . . . . . . . . . . . .

VitaminsNicotinic acid. . . . . . . . . . . . . . .Riboflavin (B2) . . . . . . . . . . . . . .Vitamin C . . . . . . . . . . . . . . . . . .

AliphaticsSorbic acid. . . . . . . . . . . . . . . . .

AntibioticsPenicillins . . . . . . . . . . . . . . . . .Erythromycins. . . . . . . . . . . . . .

Peptide hormonesBovine growth hormone. . . . . .Porcine growth hormone . . . . .Ovine growth hormone. . . . . . .

Viral antigensFoot-and-mouth disease virus.

Feed additiveFeed additiveFeed additiveFeed additive

Feed additiveFeed additiveFeed additive

Feed preservative

Feed additive, prophylacticFeed additive, prophylactic

Growth promoterGrowth promoterGrowth promoter

Vaccine

Rous sarcoma virus . . . . . . . . . . VaccineAvian Ieukemiavirus . . . . . . . . . VaccineAvian myeloblastosis virus. . . . Vaccine

EnzymesPapain . . . . . . . . . . . . . . . . . . . . . Feed additiveGl ucose oxida se . . . . . . . . . . . . Feed preservative

PesticidesMicrobial . . . . . . . . . . . . . . . . . . InsecticideAromatic. . . . . . . . . . . . . . . . . . . Insecticide

InorganicAmmonia . . . . . . . . . . . . . . . . . . Fertilizer

SOURCE: Genex Corp.

tables from 1-B-12 to I-B-32 project that within 20years all these products could be manufacturedusing genetically engineered microbial strains on a

more economical basis than using today’s conven-

tional technologies. In many cases, the time required

to apply genetically engineered strains in commercial

fermentations could be reduced to as little as 5 years.

The impact of genet ic engineering on selected

markets is shown in table I-B-33. only five product

categories are considered here, and in one, amino

acids, only a few of the compounds comprising it are

evaluated. The products represented in the five cate-

gories currently have a total market value exceeding

$800 million. However, within 20 years this market

value could rise to over $5 billion (in 1980 dol l a r s )due largely to the application of genetic engineering.

In a number of cases, the desired products would

most likely not b e available in sign ificant qu antities if

not for the application of genetic engineering tech-

nology.



Appendix l-B—A Timetable for the Commercial Production of Compounds q 279

Table l- B-7.—Chemicals: Aliphatics

Compound End use

Acetic acida. . . . . . . . . . . . . . . .

Acrylic acida. . . . . . . . . . . . . . . .Adipic acid

a. . . . . . . . . . . . . . . .

Bis (2-ethylhexyl) adipate. . . . .Citronella. . . . . . . . . . . . . . . . . .

Citronellol . . . . . . . . . . . . . . . . .Ethanol

a. . . . . . . . . . . . . . . . . . .

Ethanolamine. . . . . . . . . . . . . . .Ethylene glycol

a. . . . . . . . . . . .

Ethylene oxidea. . . . . . . . . . . . . .

Geraniol . . . . . . . . . . . . . . . . . . .Glycerol a . . . . . . . . . . . . . . . . . .Isobutylene . . . . . . . . . . . . . . . .

Itaconic acid . . . . . . . . . . . . . . .Linalool. . . . . . . . . . . . . . . . . . . .Linalyl acetate. . . . . . . . . . . . . .Methane. . . . . . . . . . . . . . . . . . .Nerol. . . . . . . . . . . . . . . . . . . . . .Pentaerythritol. . . . . . . . . . . . . .Propylene giycol

a. . . . . . . . . . .

Sorbitol. . . . . . . . . . . . . . . . . . . .

a-terpineol . . . . . . . . . . . . . . . . .a-terpinylacetate . . . . . . . . . . .

Miscellaneous acyclicMiscellaneous acyclicMisceilaneous acyclicPlasticizerFlavor/perfume material

Flavor/perfume materialMiscellaneous acylicMiscellaneous acyclicMiscellaneous acyclicMiscellaneous acyclicFlavor/perfume materialMiscellaneous acyclicMiscellaneous acyciic,

flavor/perfume materialPlastics/resinFlavor/perfume materialFlavor/perfume materialPrimary petroleum productFlavor/perfume materialMiscellaneous acyclicMiscellaneous acyiicMiscellaneous acyclic

Flavor/perfume materialFlavor/perfume material

alndicates compounds also identified by the Massachusetts Institute of

Technology. The following additional chemicals were identified by MIT asamenable to biotechnological production methods: acetaldehyde, acetoin,acetone, acety lene, acry l ic ac id, butadiene, butano~ butyl acetate,butyraldehyde, dihydroxyacetone, ethyl acetate, ethyl acrylate, ethylene, for-maldehyde, isoprene, isopropanol, methanol, methyl ethyl ketone, methylacrylate, propylene, propylene oxide, styrene, vinyl acetate.

SOURCE: Genex Corp. and the Massachusetts Institute of Technology.

Table l-B-8.—Chemicals: Aromatics andMiscellaneous


AromaticsAniline . . . . . . . . . . . . . . . . . . . . Cyclic intermediateBenzoic acid. . . . . . . . . . . . . . . . Cyclic intermediateCresols. . . . . . . . . . . . . . . . . . . . Cyclic intermediatePhenol . . . . . . . . . . . . . . . . . . . . Cyclic intermediatePhthalic anhydride . . . . . . . . . . Cyclic intermediateCinnamaldehyde. . . . . . . . . . . . Flavor/perfume materialDiisodecyl phthalate. . . . . . . . . PlasticizerDioctyle phthalate. . . . . . . . . . . Plasticizer

InorganicAmmonia . . . . . . . . . . . . . . . . . . Manufacturing processesHydrogen . . . . . . . . . . . . . . . . . . Manufacturing processes

EnzymesPepsin. . . . . . . . . . . . . . . . . . . . . Manufacturing processesBacillus protease. . . . . . . . . . . . Manufacturing processes

Mineral leachingTransition metals (cobalt, Inorganic intermediates;nickel, manganese, iron). . . . . . catalysts

Biodegradation . . . . . . . . . . . . . Removal of organicphosphates, arylsulfonates, andhaloaromatics

Table l- B-9.—Energy Products


EnzymesEthanol dehydrogenase. Manufacturing processesHydrogenate. . . . . . . . . . Manufacturing processes

Biodegradation. . . . . . . . Petroleum byproducts removal

AliphaticsMethane. . . . . . . . . . . . . . FuelEthanol . . . . . . . . . . . . . . Fuel

InorganicHydrogen. . . . . . . . . . . . . Fuel

Mineral leachingUranium. . . . . . . . . . . . . . Fuel

SOURCE: Genx Corp.

Table I- B- 10.—Total Market Values for theVarious Product Categories

Number of Current valueProduct category compounds ($ millions)

Amino acids. . . . . . . . . . . . . . . . 9 $ 1,703.0Vitamins. . . . . . . . . . . . . . . . . . . 6 667.7Enzymes. . . . . . . . . . . . . . . . . . . 11 217.7Steroid hormones . . . . . . . . . . . 6 376.8Peptide hormones. . . . . . . . . . . 9 263.7Viral antigens. . . . . . . . . . . . . . . 9 N/AShort peptides. . . . . . . . . . . . . . 2 4.4Nucleotides. . . . . . . . . . . . . . . . 2 72.0Miscellaneous proteins . . . . . . 2a 300.0Antibiotics. . . . . . . . . . . . . . . . . 4

b

4,240.0Gene preparations. . . . . . . . . . . 3 N/APesticides . . . . . . . . . . . . . . . . . 2

b

100.0Aliphatics:

Methane. . . . . . . . . . . . . . . . .12,572.0Other. . . . . . . . . . . . . . . . . . . . 24 c

2,737.5Aromatics. . . . . . . . . . . . . . . . . . 10

c

1,250.9Inorganic . . . . . . . . . . . . . . . . . 2 2,681.0Mineral leaching . . . . . . . . . . . . 5 N/ABiodegradation . . . . . . . . . . . . . N/A N/A

Totals . . . . . . . . . . . . . . . . . 107 $27,186.7d

%nly two of a number of compounds are considered here.%hese numbers refer to major classes of compounds; not actual numbers of

compounds.cThege numbers refer only to those compounds rePresenting the k9est

market volume In classes specified in the text.dcument value excluding methane = $14,614,7W,~.

SOURCE: Genex Corp.

SOURCE: Genex Corp.




Table I-B-11.-Amino Acids: Market Information

Current market data

Marketvolume Bulk cost Market value

Compound 1,000 lb $/lb ($ millions)

Arginine . . . . . . . . 900 12.73 11.46

Aspartate . . . . . . . 3,000 2.86 8.6Cysteine. . . . . . . . 600 22.75 13.6Glutamate. . . . . . . 600,000 1.80 1,080.0Lysine . . . . . . . . . 129,000 2.10 258.0Methionine. . . . . . 210,000 1.40 294.0Phertylalanine. . . . 300 38.18 11.46Threonine. . . . . . . 300 58.18 16.2Tryptophan. . . . . . 225 43.18 9.71

SOURCE: Compiled by Genex Corp. from data in references 1,2, and 3.

Table l-B-12.—Amino Acids: Technical Information

Is precursor Time to implementrenewable/non- Alternate commercial fermentation by

Typical synthetic renewable precursor by genetically engineeredCompound process Typical precursor limited fermentation strain

Arginine . . . . . . . .

Aspartate . . . . . . .

Cysteine . . . . . . . .

Glutamate. . . . . . .

Lysine. . . . . . . . . .

Methionine. . . . . .

Phenylalanine. . . .

Threonine. . . . . . .

Tryptophan. . . . . .

fermentation

fermentation

extraction

fermentation

fermentation

chemical

chemical

fermentation

chemical

glucoseand NH4

+a

fumaric acidand ammonia

proteinhydrolysis

glucoseand NH4

+

glucoseand NH4

+

B-methylmercaptoproplonaldehyde

a-acetamino-cinnamic acid

glucoseand NH4

+

a-ketoglutaricphenylhydrazone

renewable

limited

renewable

renewable

renewable

nonrenewable

limited

renewable

nonrenewable

glucoseand NH4

+

glucoseand NH4

+

glucoseand NH4

+

5 yrs.

5 yrs.

5 yrs.

5 yrs.

5 yrs.

10 yrs.

5 yrs.5 yrs.

5 yrs.

5 yrs.

aArnmonlum ion.

SOURCE: Compiled by Genex Corp. from data in references 2,3,4, and 5

Table l- B-13.-Vitamins: Market Information

Current market data



Nicotinic acid. . . . 1,400 1.82 2.5Riboflavin (B2). . . . 22 15.40 0.34Vitamin B12 . . . . . . 6,991.60 153.8Vitamin C . . . . . . . 90,0% 4.50 405.0Vitamin D . . . . . . . 12 42.50 0.51Vitamin E . . . . . . . 3,641 29.00 105.6

SOURCE: Compiled by Genex Corp. from data in references 1,6,7,8, and 9.



Appendix l-B—A Timetable for the Commercial production of Compounds . 281

Table l- B-14.—Vitamins: Technical Information



Nicotinic Acid . . . chemical aikyl a-subst. nonrenewable glucose 10 yrs.

and NH4

+a

Riboflavin (B2). . . . fermentation pyridines renewable — 10 yrs.glucose

Vitamin B12

. . . . . . fermentation carbohydrates renewable — 10 yrs.

Vitamin C . . . . . . . semisynthetic glucose or renewable 10 yrs.sorbitol

Vitamin D . . . . . . . fermentation glucose renewable glucose 10 yrs.

Vitamin E . . . . . . . extraction wheat germ oil limited glucose 15 yrs.

aAmmonium ion,

SOURCE: Compiled by Genex Corp. from data in references 4,7,8, 10,11, and 12.

Table I-B-1 5.—Enzymes: Market Information

Current market data



a-amylase. , . . . . . 600 19.33 11.6Amyloglucosidase 600Asparaginase. . . . (lnformation not available)Bacillus protease. 1,000 8.28 8.2Ethanoldehydrogenase . (Information not available)

Glucose isomerase 100 40000 400Glucose oxidase . 0.80Hydrogenate . . . . (Iformation not available)Papain. . . . . . . . . . 200 59.00 11.8

Pepsin. . . . . . . . . . 10 360.00 3.8Rennin. . . . . . . . . . 24 696.00 40.0Tyrosine

hydroxylase . . . . (Information not available)Urokinase. . . . . . . 60,000 IUa

89.5

alu = international units.




282 q Impacts of Applied Genetics— Micro-Organ/sins, Plants, and Animals

Table I- B-16.-Enzymes: Technical Information



a-amylase. . . . . . .

AmyloglucosidaseAsparaginase. . . .

Bacillus protease.

Ethanoldehydrogenase

Glucoseisomerase. . . . .

Glucose oxidase .

Hydrogenate . . . .

Papain. . . . . . . . . .

Pepsin. . . . . . . . . .Rennin. . . . . . . . . .

Tyrosine . . . . . . . .

Urokinase. . . . . . .

fermentation

fermentationextraction

fermentation

fermentation

fermentation

extraction

fermentationfermentation

extraction

extraction

molasses

molassestissue culture

molasses

(Information not available)

glucoseand NH4

+a

molasses


papaya

molassesmolasses

tissue culture

tissue culture

renewable

renewablerenewable

renewable

renewable

renewable

renewable

renewablerenewable

renewable

renewable

glucoseand NH4

+

glucoseand NH4

+

glucoseand NH4

+

glucoseand NH4

+

glucoseand NH4

+

glucose

5 yrs.

5 yrs.5 yrs.

5 yrs.

10 yrs.

5 yrs.

5 yrs.

10 yrs.

5 yrs.

5 yrs.5 yrs.

5 yrs.

5 yrs.

aArnrnonium ion.

SOURCE: Compiled by Genex Corp. from data in references 4,5,13,14,16,17, and 18.

Table l=B=17.-Sterold Hormones: Marketinformation

Current market data

Marketvolume Bulk cost Market valueCompound 1,000 lb $/lb ($ millions)

Corticoids. . . . q . . 305.8Cortisone . . . . . . . N/A 208.84 N/APrednisone. . . . . . N/A 467.62 N/APrenisolone . . . . . N/A 463.06 N/AAldosterone . . . . . N/A N/A N/A

Androgens 10,8Testosterone . . . . (Information not available)

Estrogens. . . . . . . 60.2Estradiol . . . . . . . . (Information not available)

SOURCE: Compiled by Genex Corp. from data In references 1 and 4.



Appendix l-B—A Time table for the Commercial Production of Compounds q 283

Table I-B-18.—Steroid Hormones: Technical Information



CorticoidsCortisonePrednisone. . . . . .

PredisoloneAldosterone

AndrogensTestosterone . . . .

EstrogensEstradiol . . . . . . . .

semisynthetic diosgenin or renewable glucose 10 yrs.stigmasterol

renewable glucose 10 yrs.semisynthetic chemicalmodification

of cholesterol

semisynthetic chemical renewable glucose 10 yrs.modification of

cholesterol


.

Table l- B-19.—Peptide Hormones: MarketInformation

Current market data

Compound

ACTHa. . . . . . . . . .

Bovine growthhormone. . . . . .

Endorphins. . . . . .Enkephalins . . . . .Glucagon . . . . . . .Human growth

hormone. . . . . .Insulin. . . . . . . . . .Ovine growth

hormone. . . . . .Porcine growth

hormone. . . . . .Vasopressin. . . . .

Marketvolume Bulk cost Market value1,000 lb $/lb ($ millions)

N/A N/A 5.6

0.0 0.0(lnformation not available)(Information not available)(Information not available)

N/A N/A 75.0N/A N/A 163.1

0.0 0.0 0.0

0.0 0.0(lnformation not available)

aAdrenoco~icotropic hormone.

SOURCE: Compiled by Genex Corp. from data in reference 4.




Table I- B-20.-Peptide Hormones: Technical Information


Typical synthetic renewable precursor by genetically engineered

Compound process Typical precursor limited fermentation strain

ACTHa. . . . . . . . . .

Bovine growthhormone. . . . . .

Endorphins. . . . . .

Enkephalins. . . . .

Glucagon . . . . . . .

Human growthhormone. . . . . .

Insulin. . . . . . . . . .

Ovine growth

hormone. . . . . .

Porcine growthhormone. . . . . .

Vasopressin. . . . .

extraction

extraction

extraction

extraction

extraction

extraction

extraction

extraction

extraction

extraction

adrenal cortex

anterior pituitary

brain

brain

pancreas

anterior pituitary

pancreas

anterior pituitary

anterior pituitary

posterior pituitary

limited

limited

limited

limited

limited

limited

limited

limited

limited

limited

glucoseand NH4

+b

glucoseand NH4

+

glucoseand NH4

+

glucoseand NH4

+

glucoseand NH4

+

glucoseand NH4

+

glucoseand NH4

+

glucoseand NH4+

glucoseand NH4

+

glucose,

5 yrs.

5 yrs.

5 yrs.

5 yrs.

5 yrs.

5 yrs.

5 yrs.

10 yrs.

10 yrs.

5 yrs.and NH4

+

aArjren~@iCOt@C hormone.

bAmmonium ion.

SOURCE: Compiled by Genex Corp. from data in references 4,23, and 24.

Table l- B-21.—Vjral Antigens: Market Information

Current market data



Avian leukemiavirus. . . . . . . . . . . (Information not available)

Avian myeloblastosisvirus. . . . . . . . . . . (information not available)

Epstein-Barr virus 0.0 0.0 0.0Hepatitis virus . . . 0.0 0.0 0.0Herpes virus. . . . . 0.0 0.0 0.0Hoof and mouth

disease virus . . 0.0influenza virus . . . (lnformation not available)Reoviruses . . . . . . 0.0 0.0 0.0Rous sarcoma

virus. . . . . . . . . . . (Information not available)Rubella virus. . . . . (Information not available)Varicella virus . . . (information not available)




Appendix I-B—A Timetable for the Commercial Production of Compounds q 2 8 5

Table l- B-22.—Viral Antigens: Technical Information



Avian leukemia. . .virus

Avianmyeloblastosis.virus

Epstein-Barr virus

Hepatitis. . . . . . . .viruses

Herpes . . . . . . . . .viruses

Hoof and mouth. .disease virus

Influenza. . . . . . . .viruses

Reoviruses . . . . . .

Rous sarcoma . . .virus

Rubella . . . . . . . . .virus

Varicella . . . . . . . .virus



tissue culture Iymphoblasts







tissue culture duck embryoniccells


glucoseand NH4

+a

glucoseand NH4

+

renewable glucoseand NH4

+

glucoseand NH4

+

glucoseand NH4

+

glucoseand NH4

+

glucoseand NH4

+

glucoseand NH4

+

glucoseand NH4

+

renewable glucoseand NH4

+

glucoseNH4

+

5 yrs.

5 yrs.

5 yrs.

5 yrs.

5 yrs.

5 yrs.

10 yrs.

15 yrs.

5 yrs.

5 yrs.

5 yrs.

aAmmonium ion.

SOURCE: Compiled by Genex Corp. from data in references 4 and 25.

Table l-B-23.—Short Peptides, Nucleotides, andMiscellaneous Proteins: Market Information

Current market data


Product category 1,000 lb $/lb ($ millions)

Short peptidesa

Aspartame . . . . . . 40 110.00 4.4Glycine-histidine-

Iysine. . . . . . . . . . (Information not available)

Nucleotides b

51-IMP

c. . . . . . . . . 4,000 12.00 48.0

51-GMP d . . . . . . . . 2,000 12.00 24.0

Miscellaneous proteinsa

Interferon . . . . . . . N/A N/A 50.0Human serumalbumin. . . . . . . . 250 1,000.00 250.0

Monoclinal

antibodies. . . . . . (Information not available)aData from references 4 and 26.

bData from references 4 and 27.

c5’-inosinic acid.d5’-guanylic acid.

‘Data from reference 4.

SOURCE: Compiled by Genex Corp.



286 q Impacts of Applied Genet/cs—Micro-Organisms, Plants, and Animals

Table l- B-24.—Short Peptides, Nucleotides, and Miscellaneous Proteins: Technical information


Typical synthetic renewable precursor by genetically engineeredProduct category process Typical precursor limited fermentation strain

Short peptidesa

Aspartame . . . . . . chemical phenylalanine & renewable glucose 5 yrs.

aspartic acid and NH4+

Glycine-histidine-Iysine. . . . . . . . . . . extraction human serum renewable glucose 5 yrs.

and NH4+

Nucleotides c

51-lMP

d. . . . . . . . . extraction yeast renewable glucose 10 yrs.

and NH4+

51-GMPe. . . . . . . . . extraction yeast renewable glucose 10 yrs.and NH4

+

Mlscellaneous proteinsf

Interferon . . . . . . . extraction or leukocytes, renewable glucose 5 yrs.tissue culture Iymphoblasts, and NH4

+

or fibroblasts

Human serumalbumin . . . . . . . . . extraction human serum renewable glucose 5 yrs.

and NH4+

Monoclinalantibodies . . . . . somatic cell various cells renewable glucose 10 yrs.

hybridization and NH4+

%ata from reference 4 and 27.bAmmonium ion.cData from references 4 and ~.%’-inoalnic acid.‘5’-guanyiic acid.‘Data from reference 4.

SOURCE: Compiied by Genex Corp.

Table I- B-25.-Antibiotics, Gene Preparations, andPesticides: Market information

Current market data


Product category 1,000 lb $/Ib ($ millions)

Antibiotic@Penicillins. . . . . . . 49,300 22.11 1,080.0Tetracycline. . . . 29,300 34.13 1,000.OCephalosporins. . 4,210 114.00 460.0Erythromycins . . . (Information not available)

Gene preperationsb

Sickle cell anemia 0.0 0.0 0.0Hemophilias. . . . . 0.0Thallasemias . . . . 0.0 0.0 0.0

Pesticidesc

Microbial. . . . . . . . N/A N/A 25.0

Aromatics . . . . . . . N/A N/A 75.0aD

at

afrom references 4,28, and 9.

bData from references 4 and X.

cData from references 4 and ~.




Appendix I-B—A Timetable for the Commercial Production of Compounds q 287

Table l- B-26.—Antibiotics, Gene Preparations, and Pesticides: Technical Information



AntibioticsPenicillins. . . . .

Tetracycline . .

Cephalosporins

Erythromycins .

. .

. .

. .

Gene preprationsb

Sickle cell anemia

Hemophilias. . . . .

Thallasemias . . . .

Pestlcidesc

Microbial. . . . . . . .

Aromatics. . . . . . .

fermentation lactose &

semisynthetic nitrogenous oilsfermentation lactose &

nitrogenous oils

fermentation lactose &nitrogenous oils

fermentation lactose &nitrogenous oils

(No process existscurrently)



fermentation molasses &fishmeal

semi synthetic naphthalene

limited

limited —

limited

limited

glucoseand NH4

+d

glucoseand NH4

+

glucoseand NH4

+

renewable

n o n r e n e w a b l e —

10 yrs.

10 yrs.

10 yrs.

10 yrs.

15 yrs.

20 yrs.

20 yrs.

5 yrs.

10 yrs.

Coata from references 4 and ~.Data from references 4,5,28, 3~ t an d 32 .

bData from reference 4. ‘Ammonium ion.


Table l= B-27. -AIiphatics: Market Information

Current market data



Acetic acid . . . . . . 823,274Acrylic acid. . . . . . 46,503

Adipic acid . . . . . . 181,097Bis (2-ethylehexyl) 43,015

adipateCitronella . . . . . . . 394Citronellol. . . . . . . 1,443Ethanol . . . . . . . . . 1,048,000Ethanolamine. . . . 320,236Ethylene glycol . . 3,137,000Ethylene oxide . . . 525,113Geraniol . . . . . . . . 2,307Glycerol . . . . . . . . 116,612Isobutylene. . . . . . 597,712Itaconic acid. . . . . 200Linalool. . . . . . . . . 3,341Linalyl acetate . . . 1,535Methane . . . . . . . . 878,000,000Nerol. . . . . . . . . . . 462

Pentaerythritol. . . 117,085Propionic acid . . . 62,848Propylene glycol . 525,527Sorbic acid . . . . . . 20,000Sorbitol. . . . . . . . . 160,267a-terpineol . . . . . . 2,416a-terpinyl acetate. 1,066

0.230.430.500.49

3.904.500.240.460.31

3.250.540.950.832.603.50

0.0134.20

0.620.210.732.150.361.281.30

189.420.0

90.521.1

6.5251.5147.3972.5189.0

7.563.0

567.20.28.75.4

11,573.01.9

72.613.2

173.443.057.7

3.01.4

SOURCE: Compiled by Genex Corp. from data in references 1,4,9, and 33.




Table l- B-28.—Aliphatics: Technical Information



b

Acetic acid. . . . . . . . .

Acrylic acid . . . . . . . .Adipic acid. . . . . . . . .Bis (2-ethylhexyl). . . .

adipateCitronella . . . . . . . . .Citronellol . . . . . . . . .Ethanol. . . . . . . . . . . .Ethanolamine . . . . . .Ethylene glycol . . . . .Ethylene oxide. . . . . .Geraniol . . . . . . . . . . .Glycerol . . . . . . . . . . .Isobutylene . . . . . . . .Itaconic acid . . . . . . .Linalool . . . . . . . . . . .Linalyl acetate . . . . . .Methane. . . . . . . . . . .

Nerol. . . . . . . . . . . . . .Pentaerythritol . . . . .

Propionic acid . . . . . .

Propylene glycol . . . .Sorbic acid. . . . . . . . .

Sorbitol. . . . . . . . . . . .a-terpineol . . . . . . . . .a-terpinyl acetate . . .

chemical

chemicalchemicalchemical

chemicalchemicalchemicalchemicalchemicalchemicalchemicalchemicalchemical

fermentationchemicalchemicalchemical

chemicalchemical

chemical

chemicalchemical

chemicalchemicalchemical

methanolor ethanol

ethylenephenolphenol

isobutyleneisobutylene

ethyleneethyleneethyleneethylene

isobutylenesoap manuf.petroleummolasses

isobutyleneisobutylenenatural gas

isobutyleneacetaldehyde &formaldehyde

ethanol &carbon monoxide

propylenecrotonaldehyde &

malonic acidglucose

isobutyleneisobutylene

nonrenewable

nonrenewablenonrenewablenonrenewable

nonrenewablenonrenewablenonrenewablenonrenewablenonrenewablenonrenewablenonrenewablenonrenewablenonrenewable

renewablenonrenewablenonrenewablenonrenewable

nonrenewablenonrenewable

limited

nonrenewablenonrenewable

renewablenonrenewablenonrenewable

glucose

glucoseglucoseglucose

glucoseglucoseglucoseglucoseglucoseglucoseglucoseglucoseglucose

. . .

glucoseglucosesewage

glucoseglucose

glucose

glucoseglucose

glucoseglucose

10 yrs.

10 yrs.10 yrs.

20 yrs.

20 yrs.20 yrs.5 yrs.10 yrs.5 yrs.5 yrs.

20 yrs.5 yrs.10 yrs.5 yrs.

20 yrs.20 yrs.10 yrs.

20 yrs.10 yrs.

10 yrs.

10 yrs.15 yrs.

10 yrs.20 yrs.20 yrs.

%Vherever glucose ia mentioned, other carbohydrates maybe aubatituted, including starch and cellulose.bln many Casea these times are bgsed on more readily developed fermentations using nonrenewable or limited hydrocarbons as Precursors.

SOURCE: Compiied by Genex Corp. from data in references 4,33,34, and 35.

Table I= B-29.-Aromatics: Market Information

Current market data



Aniline . . . . . . . . . . . . 187,767 0.42 78.9Aspirin . . . . . . . . . . . . 32,247 1.41 45.5Benzoic acid . . . . . . . 36,822 0.47 17.3Cinnamaldehyde. . . . 1,098 2.10 3.4Cresols. . . . . . . . . . . . 94,932 0.54 51.2Diisodecyl

phthalate . . . . . . . . 151,319 0.42 63.6Dioctyl phthalate. . . . 391,131 0.42 164.3p-acetaminophenol. . 20,000 2.65 53.0

Phenol . . . . . . . . . . . . 1,431,000 0.36 515.2Phthalic anhydride . . 646,289 0.40 258.5




Appendix l-B—A Timetable for the Commercial Production of Compounds q 289

Table l- B-30.-Aromatics: Technical Information

Is precursorrenewable/non-

Typical synthetic renewableCompound process Typical precursor limited

Aniline . . . . . . . . . . . . chemical benzene nonrenewable

Aspirin . . . . . . . . . . . . chemical phenol nonrenewableBenzoic acid . . . . . . . chemical tar oil nonrenewableCinnamaidehyde. . . . chemical benzaldehyde nonrenewable

acetaldehyde

Cresols. . . . . . . . . . . . chemical phthalic nonrenewablean hydride

Diisodecyl . . . . . . . . . chemical coal tar nonrenewablephthalate

Dioctyl . . . . . . . . . . . . chemical coal tar nonrenewablephthalate

p-acetaminophenol. . chemical nitrobenzene nonrenewable

Phenol . . . . . . . . . . . . chemical coal tar nonrenewablePhthalic . . . . . . . . . . . chemical coal tar nonrenewable

anhydride

Time to implementAlternate commercial fermentation by

precursor by genetically engineeredfermentation strain

aromatica

10 yrs.

aromatic 5 yrs.aromatic 10 yrs.aromatic 20 yrs.

aromatic 10 yrs.

aromatic 20 yrs.

aromatic 20 yrs.

aromatic 5 yrs.aromatic 10 yrs.aromatic 15 yrs.

aAromatic refers t. benzene or benzene derivative. Eventually it is anticipated that Iignin, a renewable resource, would sewe as a Precursor.


Table I-B-31 .—lnorganics and Mineral Leaching:Market Information

Current market data

Marketvolume Bulk cost Market va l ue

Product category 1,000 lb $/lb ($ millions)

InorganicAmmonia . . . . . . . 33,400,000 0.06 2,004.0Hydrogen . . . . . . . 451,000 0.15 677.0

Mineral leachingUranium . . . . . . . . (Information not available)Transition metals. (Information not avaiiable)

(cobalt, nickel, manganese, iron)


Table l- B-32.-lnorganics and Mineral Leaching: Technical Information



InorganicAmmonia . . . . . . . chemical water and coke nonrenewable nitrogen(air) 15 yrs.Hydrogen . . . . . . . catalytic petroleum nonrenewable water and air 15 yrs.

reforming

Mineral leaching Uranium . . . . . . . . (Information not available)Transition metals. (Information not available)

(cobalt, nickel, manganese, iron)




Appendix I-C

C h e m ic a l a n d B io lo gic a l P r o ce s se s

A comparison was made of waste stream pollution

for chemical and biological processes. Ideally, the

comparison should be between the two processes

used in the production of the same end product.Since such data do not current ly exist at the in-

dustrial level, the comparison was made between the

chemical production of a mixture of chemicals and

the biological production of alcohol and antibiotics.one noteworthy parameter is the 5-day biochemical

oxygen demand (BOD5) —the oxygen required over a5-day period by organisms that consume degradable

organics in the waste stream. If the oxygen demand

is too high, the discharge of such a stream into abody of water will deplete the dissolved oxygen to

the point that it threatens aquatic life. An important

variable that must be considered along with the BOD

is the CO D (the chemical oxygen d emand ). Large dif-

ferences between the COD and BOD of a waste sys-tem can indicate the presence of nonbiodegradable

su b stan ces. A l th o u g h t h e c on v e n t io n a l p r o ce s s

stream shown in table 1-C-1 has less BOD5 than the

biological process stream, its COD content probably

means that non biodegradables are present , and

specialized waste treatment is necessary.BOD is one area where traditional fermentation

based processes have posed pollut ion problems.

Batch fermentation processes typically generatelarge quantities of dead cells and residual nutrients

that cause a large BOD if they are dumped directly

into a dyn amic aquatic environm ent. (See table I-C-1.)

This difficulty can be circumvented by the use of

spent cell material as an animal feed supplement or

Table I-C-1.— Waste Stream Pollution Parameters:Current Processes v. Biological Processes

Compounds: Mixed chemicals, including ethylene

oxide, propylene oxide, glycols, amines, and ethers

Current BiologicalPollution parameters processes processes

Alkalinity (mg/l).

BOD5 a(mg/l). . .

Chlorides (mg/l)C O Db(mg/l). . . .

Oils (mg/l) . . . . .

pH . . . . . . . . . . .

Sulfates (mg/l) .

. . . . . . . . . . 4,060 0

. . . . . . . . . . 1,950 4,000-12,000

. . . . . . . . . . 430-800 0

. . . . . . . . . . 7 ,970-8540 5,000-13,000

. . . . . . . . . . 547 0

. . . . . . . . . . 9.4-9.8 4-7

. . . . . . . . . . 655 0Total nitrogen (mg/l). . . . . . . 1,160-1,253 50-200Phosphates (mg/l) . . . . . . . . 0 50-200

as.day b iOk@Ca l oxygen demand.

bchemical oxygen demand.


as a fertilizer. These applications have been inten-

sively investigated and have met w ith success.

Because of the renewed interest generated by the

potentials of genetic engineering, some traditionalfermentation systems are being redesigned. Immo-bilization allows the reuse of cells that would other-

wise be discarded. These systems can be used con-

tinuously for several months as compared with theusual fermentation time in a batch process of about

one week or less. [remobilized operations create

waste cells much less often than batch systems, and

therefore generate less BOD.

292



Appendix I-D

T h e I m p a c t o f G e n e t i c s o n E t h a n o l —

A C as e S t udy

Objec t ive

This study examines how genetics can and will af-

fect the utilization of biomass for liquid fuels produc-

tion. There are two major areas where genetics are

applicable. One is in plant breeding to improve avail-

ability (both quantity and quality) of biomass re-

sources (with existing and previously unused land);

the second is in the application of both classical

mutation and selection procedures and the new ge-

netic engineering techniques to develop more effi-

cient microbial strains for biomass conversion. Ex-

amples of goals in a plant breeding program would

include improvements in photosynthetic efficiencies,

increased carbohydrate content, decreased or modi-fied lignin content, adaptation of high productivity

Figure l-D-l .—An Overview of Alternative

plants to poor quality land, improved disease resist-ance, and so forth. However, the focus here is entire-

ly on the second area, the use of genetics to improve

microbial-based conversion to produce ethanol.

In order to assess the type and extent of im-provements in micro-organisms that might benefitethanol produ ction, its process technology andeconom ics mu st first be examined . An overview of the biomass conversion technology is presented infigure I-D-1; processes are defined mainly on thebasis of the primary raw material and th e type of pretreatment required to produce mono- or di-saccharides prior to fermentation. In addition, thereare several alternative fermentation routes to pro-

duce ethanol; these are characterized by the type of micro-organisms and will be examined with the in-

Routes for Conversion of Biomass to Ethanol


76-565 0 - 81 - 20

293




tent of quantifying the potential impact of genetic im-provement on each one. It is interesting to note thateach type of organism has its substrate restrictions,

.

and only the anaerobic bacteria such as Clostridiumthermosaccharolyticum and C. thermohydrosulfori-cum can utilize all of the available sub strate.

Subs t ra te p re t re a tme n t

Pretreatment refers to the processing that is re-

quired to convert a raw material such as sugarcane,starch, or cellulosic biomass to a product that is

fermentable to ethanol. In most cases, the pretreat-ment is either extraction of a sugar or hydrolysis of a

polysaccharide to yield a mono-or disaccharide.

EXTRACTION OF SUGAR

Sugar crops such as sugarcane, sugar beets, or

sweet sorghum are highly desirable raw materials

for producing ethanol . These crops contain high

amounts of sugars as sucrose. In addition, the yield

of fermentable material per acre is high; sugarcaneand sugar beets yield 7.5 and 4.1 dry tons of biomass

per acre, respectively. ’

Sugar is extracted from cane or beets with hotwater and then recrystallized. The resulting sugars

are utilized directly by organisms having invertase

activity (to split sucrose to glucose plus fructose).

Molasses, a sugary byproduct of the crystallization of

sucrose, may also contain sucrose although in most

cases it is inverted with acid.

The primary use for sugar crops is food sugar.

Sugar sells for over 20 cents/lb. Molasses, which cur-

rently sells for about $100/ton (about 10 cents/lb

sugar) is used extensively as an animal feed. Substan-

t ial amounts of both sugar and molasses are im-ported into the United States for food uses and are

therefore unavailable for ethanol production, There

are proposals to increase sugar production for use as

an energy c rop; however , th i s wi l l requi re the

development of new land for sugar produ ction.

STARCH

The primary raw material for ethanol fermenta-

tion in the United States is cornstarch. Corn proc-

essed by wet milling, yields about 36 lb of starch

from each 56 lb bu; this amount of starch will pro-

duce 2.5 gal of absolute ethanol. Corn yields are

typically 80 to 120 bu/acre so that 200 to 300 gal of

ethanol can be derived per acre of corn per year.Pretreatment of starch is initiated by a gelatiniza-

tion step whereby a starch slurry is heated for 5 min

at 105° C. After cooling to 98° C, œ-amylase is added

‘Paul B. Weisz and John F. Marshall, Science 206:24, 1979.

to break down the starch to about 15DE (dextrose

equivalents). This process of liquefaction reduces the

viscosity such that the solution can be easily mixed.

After further cooling to 30° C, glucoamylase is added

along with a starting culture of yeast so that saccha-

rification and fermentation proceed simultaneously.

The resulting fermentation, to produce typically 8 to

10 percent ethanol (volume per volume), requires 42

to 48 hr for completion. This compares with a 16- to

20-hr fermentation if sugar as molasses or cane juiceis used as the substrate. Thus, the use of starch re-

quires the addition of enzymes prior to and during

fermentation, as well as large fermenter capacity as a

consequence of the slower fermentation time com-pared with sugar substrates.

Improvement in the economy of ethanol fermenta-

t ion based on starch is possible by developing a

micro-organism tha t can produce wamylase an dglucoamylase and thus eliminate the need to add

these enzymes. Since the rate of fermentation de-

pends on the rate of starch hydrolysis, increased lev-

els of glucoamylase may enhance the rate of starch

hydrolysis and thus increase the rate of ethanol pro-

duction, This would lower the capital requirements

as well as the cost of enzyme add ition.

CELLULOSIC BIOMASS

Processes for the utilization of cellulosic biomass

to produce liquid fuels all have three features in com-

mon:

1. They employ some means of pretreatment to at

2.

3.

least effect some initial size reduction and, more

often, cause a disassociation of lignin and cellu-

lose;they involve either acid or enzymatic hydrolysis

of the cellulose and hemicellulose to producemono- and disaccharides; and

they employ fermentation to produce ethanol or

some other chemical.

A wide variety of process schemes have been pro-

posed for the conversion of cellulosic biomass to

liquid fuels; a summary of the major steps in two

a c i d hydro l ys i s a nd t h re e e nz yma t i c hydro l ys i s

schemes in shown in figures I-D-2 and I-D-3. The in-i t i a l s ize reduc t ion i s requi red to increase the

amount of biomass surface area that can be con-

t a c t e d w i t h a c i d , s o l v e n t , s t e a m , e n z y m e s , o r

chemicals that might be used to disassociate the

c e l l u l o s e a n d h e m i c e l l u l o s e f r o m t he l i gn i n .P r e t r e a t m e n t s t h a t h a v e b e e n i n v e s t i g a t e d t o

fac i l i t a te the process a re summarized in t able

I-D-1. The problems with pretreatment are that they

require energy, equipment , and often chemicals;

they result in an irretrievable loss of sugar, and in

undesirable side-react ions and byproduct forma-



Appendix l-D—The Impact of Genetics on Ethanol—A Case Study q 295

Figure l-D-2.–Alternative Schemes for Acid Hydrolysis of Cellulosic Biomass for Ethanol Production


tion. Furthermore, if acids, alkali, or organic chem-

icals are used, they must be recycled to minimize

cost or disposed of in order to prevent pollution.

In starch processing, prior to ethanol fermenta-tion, mechanical grinding, steam, and enzymes are

employed. The energy requirements are small and

contribute relatively little to the final ethanol cost.

The objective in the development of cellulose-based

processes should be to minimize both energy and

chemical requirements. The development and scale-

up of effective pretreatment technology are under

active investigation and require continued financial

support to better develop several alternative routes.

The most promising routes are: steam treatment, sol-

vent delignification, dilute acid, cellulose dissolution,

and direct fermentation.

Several different acid hydrolysis schemes have

been proposed. However, most appear as in f low

scheme A or B in figure I-D-2. Dilute acid is used to

hydrolyze the hemicellulose to pentose sugars pri-

marily and then stronger acid at higher tempera-

‘Proceedings of 3rd Annual Biomass Energv System Conference, National

Technical Information Set-vice, SfXl~rP-33-285, 1979.

tures is used to cause cellulose hydrolysis (scheme

A). A major problem with this approach is the irre-versible loss of sugars to undesirable side-product

formation. After separation of residual solids (mostly

lignin), which can be burned to provide energy for

distillation, the sugar solution is fermented by yeast

to ethanol. The pentose sugars also can be fer-

mented , bu t by organisms o ther than the e thanol

producing yeast, to other chemicals, some of which

could be used as fuels (e.g., ethanol, acetic acid,

acetone, butanol, 2,3-butanediol, etc.).

An altern ative (schem e B, figure I-D-2) to the ab ove

is to use a solvent, after pentose sugar removal, to

dissolve the cellulose, allowing its separation from

lignin. This cellulose solution is easily and efficiently

hydrolyzed to sugars. The advantage of this ap-

proach over the direct acid hydrolysis is that theyield of sugar is much higher. In the harsh acid hy-

drolysis, considerable sugar is destroyed. However,

the major disadvantage of both these schemes is that

they require recycl ing or d isposal o f ac ids and

solvents. A second problem is that almost nothing is

known about how to scale-up some of the newly de-




Figure I-D-3. —Alternative Schemes for Enzymatic Hydrolysis of Cellulosic Biomass for Ethanol Production


Table I- D-1 .-Alternative Pretreatment Methods forLignocellulose Materials

Chemical m ethods Physical methods

Sodium hydroxide (alkali) SteamAmmonia Grinding and millingChemical pulping IrradiationAmmonium bisulfite FreezingSulfiteSodium chloriteOrganic solventsAcids


veloped technology, suc h a s t ha t de ve l ope d by

groups working at Purdue University, New York Uni-

versity, and Dartmouth College. There are several

engineering problems involving both heat and mass

transfer and acid/solvent recycle that need to be eval-

uated at larger scale. At least some of this work will

be done at the process development unit now being

buil t a t the Georgia Inst i tute of Technology. The

most promising direct ions that need development

are:

q the scale-up of high rates and high yield labora-

tory hydrolysis systems, and

q the development of methods for acid and chem-

ical recycle schemes.

There are three types of approaches that have

been employed for enzymatic hydrolysis of cellulosic

biomass. These are summarized in figure I-D-3. They

all involve some initial size reduction to increase the

surface a rea ava i lable for enzymat ic a t t ack. In

schemes A and B, the incoming cellulosic biomass is

split into two streams; one is used to grow organisms

that produce cellulolytic enzymes called cellulases,

and the other is used to produ ce sugar.

In scheme A, the cellulases are recovered and then

added to a separate enzyme hydrolysis react ion.

They hydrolyze both the cellulose and hemicellulose,

and the resulting sugar solution is then passed to an

ethanol fermentation stage where hexoses are con-

verted by a yeast fermentation to ethanol. Utilization

of the pentose requires a separate fermentation. Re-




sidual lignin, which is removed before (by solvents

extraction) or after hydrolysis, is used to provide

energy for ethanol recovery, Extensive work on this

approach has been done at the University of Califor-

nia, Berkeley, and the U.S. Army Natick Laboratories.

In scheme B, the cellulase is not recovered but

rather, the whole fermentation broth from cellulase

production is added to the cellulosic biomass along

with ethanol-producing yeast. The result is a simul-

taneous cellulose hydrolysis (saccharification) and

fermentat ion. (In the production of ethanol from

starch, the starch hydrolyzing enzymes are added at

the same time as the yeast for simultaneous sacchari-

fication and fermentation.) This technology has been

demonstrated by the Gulf Oil Co. After fermentation,

the ethanol is recovered and the residual lignin can

again be used for energy for distillation. The prob-

lem of unused pentose sugar still remains and will re-

quire a separate fermentation step.

A third alternative (scheme C, figure I-D-3) shows a

simpler approach, namely a direct fermentation on

cellulose. This approach has been developed at the

Massachusetts Institute of Technology. It utilizes

bacteria that will produce cellulase to hydrolyze the

cel lulose and hemicel lulose and ferment both the

hexose and pentose sugars to ethanol in a single-

stage reactor. The advantage of this approach is a

minimal requirement for pretreatment, a combined

enzyme production, cellulose hydrolysis and ethanolfermentation, and simultaneous conversion of both

pentose and hexose sugars to ethanol. This conceptis new and work still needs to be done to increase the

ethanol concentration, minimize side product forma-

t ion, and increase the rate of ethanol production.Again, residual lignin will be used to provide the

energy for ethanol distillation.

FERMENTATION O F ETHANO L

An examination of the economics for ethanol pro-

duction shows that the dominant cost is the process

raw m aterial. As seen in table 1-D-2 the feedstock rep-

resents 60 to 70 percent of the manufacturing cost.

Thus, it is clear that any improvement in substrate

utilization efficiency is of substantial benefit. The

theoretical yields of ethanol from glucose, sucrose,

and starch or cellulose are 0.51, 0.54 and 0.57 gram

(g) ethanol/g material, respectively; the differencesresult from the addition of a molecule of water on

hydrolysis. There are several approaches to improve

the yield above the typical value of 90 to 95 percentcurrentl y achieved. These are:

. increase the ratio of ethanol produced per unit

weight of cel ls , e .g. , through cel l recycle,

vacuum fermentation, immobilized cells, or im-provement in specific productivity (g ethanol/g

m

Table I-D-2.—A Comparison of the Distribution ofManufacturing Costs for Several Ethanol

Production Processes

GrainSubstrate Molasses Corn Sorghum

Cost component (%)

Capital. . . . . . . . . . . . . . . . 9Operating . . . . . . . . . . . . . 20Feed stock. . . . . . . . . . . . . 71

Total . . . . . . . . . . . . . . . 100Cost on energy basis

($MMBtu) . . . . . . . . . . . . . 12.5Cost/gal ethanol ($/gal) . . . . 1.05Capital investment

($/ann ua l gal ) . . . . . . . . . . 1.02

122662

103060

1 0 0

14.91.25

1.05

100

12.71.07

1.75

SOURCE: “Comparative Economic Assessment of Ethanol From Biomass,”Mitre Corp., report HCP/ET-2854).

cell hr), by increasing the content and/or activi-

ty of those enzymes in the pathway to ethanol;

increase the utilization of other materials in the

substrate, e.g., the use of oligosaccharides, espe-

cially branched, in starch, and the use of con-

taminating sugars such as galactose or mannose

for hemicellulose; and

develop a route for the utilization of pentose su-

gars, especially xylose, present in hemicellulose.

The potential effect of oligosaccharides or con-

taminating sugar utilization is relatively small, since

they represent typically 1 to 3 percent of the total

sugar content. However, if cellulosic biomass con-

taining 15 to 25 percent hemicellulose is used, then

the impact of pentose conversion to ethanol is great.Cellulosic biomass is made up primarily of cellu-

lose, hemicellulose (mostly xylan) and lignin. Othercomponents such as protein, ash, fats, etc., typically

comprise about 10 percent. The composition of b io-

mass can be expressed in terms of the fol lowing

equation:

F,. + F,, + F,,= 10 [1]

1 – 1

where FC, FH, F~ ) and FA are the weight fractions of

cellulose, hemicellulose, lignin, and ash, respectively.

Assuming that the ash is 10 percent (FA = 0.1) and

that FC and FH are the only fermentable components

in the biomass, then:

F C = FH = 0.9 – F L (2)

The maximum amount of ethanol from one unit of biomass (Y~~,~) is;

YWCFC = YE/HFH = Y~jB (3)

Where Yi,,C an d Yk;lH are the yield of ethanol for cel-

lulose and hemicellulose, respectively. Equation 2

can be rearranged to relate the fractions of cellulose:




F c = 0.9 – F L - FH (4)

Substituting this into equation 3 gives:Y

E/ B+ Y

E/ c(0.9 – F

L– F

H) + Y

E/ HF

H (5)

From equation 5, the effect can be calculated of hemicellulose content and conversion yield on theoverall conversion of biomass to ethanol. Assuming alignin content of 15 percent (F

L

= 0.15) and using YE/ c= 0.57 g/ g the following equation is obtained:

YE / B

= 0.43 + F H( YE/H - 0.57) (6)

The theoretical yield value on hemicellulose,YE/H, is not well-defined because so little is kn ownabout the biochemistry of anaerobic pentosemetabolism. If one mole of ethanol is produ ced p ermole of xylose, the yield is 0.3 g ethanol/g xylose. Ittwo moles of ethanol could be obained, YE/H wouldbe 0.61; however, neither the mechanism nor thethermodynamics of the conversion is sufficientlywell-defined to allow one to expect this value. Themaximum observed values are about 0.41 gethanol/g xylose.

3The sensitivity of the overall

yield to this value is shown in figure I-D-4. The im-pact of pentose utilization depends on the amount

%. D. Wang and [:. Cooney, Massachusetts Institute of Technolo~v, un-

published results.

Figure l= D-4.-Effect of Pentose Yieid (YE/H) onOverall Yield of Ethanol from Cellulosic Biomass

(YE/B) with Varying Fractions of Hemicellulose (FH).

0.5

0 0.2 0.4 0.6

Yield on pentose (g ethanol/g pentose)


of hemicellulose present. From the value in figureI-D-4 and the observation that 70 percent of themanufacturing cost is the raw material cost, it ispossible to estimate the economic benefit of pen-tose u tilization. Equation 7 relates the overallethanol yield to the manufacturing cost:

C E = C B x 6 . 6

(7)Y E / B().7

where CE is the manufacturing cost per gallon of ethanol, CBis biomass cost (cents/ lb), 6.6 is the con-version from pou nd to gallon of ethanol, and 0.7 isthe 70-percent factor for relative biomass cost toethanol cost. For a biomass costing 2 cents/ lb andcontaining 20 percent hemicellulose, the manufac-turing cost is reduced from 59 to 43 cents/ gal, whenthe yield on pentose goes from zero to 0.6.

At the present time, there are few organisms thatproduce more than one mole of ethanol per mole of pentose and none of the usual alcohol producing

yeasts will ferment pentoses to ethanol. Addition or

improvement of the ability to use pentose will have amajor impact on the economics of ethanol produc-

tion.

The second major cost in ethanol production re-

lates to the cost of operation. Typically, 20 to 30 per-

cent of the final manufacturing cost is accounted for

by the sum of labor, plant overhead, administration,

chemical supplies, and fuel costs. The chemical sup-

plies represent less than 1 cent/gal ethanol and may

be neglected. The labor, overhead, and marketing

costs vary with plant size, but represent 11 to 7

cents/gal for a 20 to 100 million gal/yr plant, respec-

tively. Any improvement in the reduction of plant

size or complexity will reduce this cost; however, the

economic impact is small. The major component of the operating cost is the fuel charge for plant opera-

tion and for distillation. Plant operations, e.g., mix-

ing, pumping, sterilization, starch gelatinization,biomass grinding, etc., represent about 20 to 30 per-

cent of the energy cost. The remainder is for ethanol

distillation and residual solids drying. Considerable

effort has been focused on methods to improve the

energy efficiency, of distillation to reduce it from the

160)000 Btu/gal required for beverage alcohol. While

considerable differences in opinion exist as to the

minimum, a reasonable expectation is about 40,000

Btu/gal although current technology requires 69,000

Btu/gal.4Forty thousand Btu is about half of the ener-

gy content of ethanol per gallon.A discussion of process improvements relating to

ethanol recovery has two components. The first is

4Report of (he Gasohol Stud-y C,roup of the Energy Research Advisory

Soard, Department of Energv, Washington, D. C., 1980. ‘M. Gibbs, and R,

D. DeMoss, “Ethanol Formation in Pseudomonas /indneri, ” Arc/I. Biochem

Biophys. 34:478-479, 1951.



Appendix l-D—The Impact of Genetics on Ethanol—A Case Study . 299

related to operating costs and the second is related to

energy efficiency. If coal is used to provide energyfor distillation, and it is valued at $30/ton, with

10,500 Btu/lb or $1.50/million Btu, then the energy

cost for distillation (optimistically assuming 40,000

Btu/gal) is $6/gal. If lignin from cellulosic biomass is

used as a fuel, the cost is reduced further. On theother hand, if oil at $40/bbl (130,000 Btu/gal and 42

gal/bbl) or $7/million Btu is used, then the energy

cost is 28 cents/gal of ethanol.

From a common sense, economic, and political

point of view, it does not seem reasonable to utilize

l iquid fuel to produce l iquid fuel from biomass.

Therefore, it will be assumed that petroleum will not

be used for distillation and that either coal or bio-

mass will be employed.In order to assess the impact of process improve-

ments on the energy demand, it is necessary to look

at an overall material balance. This is summarized in

figure 1-D-5. Only a portion of the entering biomass

feedstock is fermented to ethanol and there are two

product st reams, one containing ethanol and the

other solids, both must be separated from water. It is

important to note that as the ethanol concentration is

increased, the energy requirement for both ethanol

recovery from the water and for drying wil l de-

crease. Therefore, the impact of developing ethanol

tolerant micro-organisms is seen as a reduction in

energy cost.

Figure l-D-5. - Process Schematic for Material andEnergy Balance

Biomass

Process heat

Ethanol

Solids


The third major cost for ethanol manufacturing is

the capital investment, which represents about 4 to

12 percent of the manufacturing cost. The capital in-

vestment is determined by the complexity of the

processes and the volumetric productivity of ethanol

production. Thus, the development of a micro-orga-nism that will require a minimum amount of feed-

stock pretreatment and wil l produce ethanol at ahigher rate will reduce the net capital investment.

The volumetric productivity (Q E) for ethanol pro-

du ction is given by:

Q E= q

pX

where q P is the specific p rodu ctivity expressed in g

ethanol per g cell hr, and X is the culture density.

Therefore, there are two approaches to obtain high

productivity: first, to choose or create an organism

with a high specific rate of ethanol production and

second, to design a process with high cell density.

The application of genetics can be used to enhance

the intracel lular enzyme act ivi ty of the enzymes

used for ethanol production. The resulting increasein q p , will result in reduced capital investment re-

quirements.There are four types of ethanol processes based

on different organisms; they are:

1. Saccharomyces cerevisiae and related yeast,

2. Saccharomyces cerevisiae/Trichoderma reesei,

3. Zymomonas mobilis, an d

4. Clostridium thermocellum/thermosaccharolyti-cum, or thermohydrosulfuricum.

The first is the traditional yeast based process using

S. cerevisiae to ferment soluble hexose sugar to eth-

anol. In the second, the substrate range is extendedto cellulose by the use of cellulase produced by T .

reesei. The third approach utilizes Z. mobilis; th i sorganism is a particularly fast and high ethanol yield-

ing one. Its range of fermentable substrates, how-

ever, is limited to solub le hexose sugars.

In many tropical areas of the Americas, Africa,

and Asia, alcoholic beverages prepared from a mixed

fermentation of plant steeps are popular. Bacteria

from the genus Zymomonas are commonly em-

ployed. In the early 1950’s, the genus Zymononas ac-

quired a certain fame among biochemists by the dis-covery that the anaerobic catabolism of glucosefollows the Enter-Doudoroff mechanism.’ This wasvery surprising, since Zymomonas was the first ex-ample of an anaerobic organism using a pathway

mainly in strictly aerobic bateria.6

In spite of its extensive use in many parts of the

world, its great social implications as an ethanol pro-

‘M. Gibbs and R. D, de Moss, “Ethanol Formation, in Psuedornonas

Iindneri,” Arch. Biochem. Biophys., 34;478-$79, 1951.

6J. Swings and J. Del . ey , “-rhe Biologv of Zymomonas, ” Bacteriological Re-views 41: 1-46, 1977.



300 q Impacts of Applied Genetics—M icro-Organisms, Plants, and Animals

du cer, and its u niq ue biochem ical position, Zymo-monas has not been studied extensively. T

The organism most often studied is Zymomonas mobilis, which can produce up to 1.9 moles of

ethanol per mole of glucose. Recent studies reported

from Australia, have established the Z. mobilis can

ferment high concentrat ions of glucose rapidly to

ethanol in both batch and continuous culture withhigher specific glucose uptakes rates for glucose andethanol production rates than for yeasts current ly

used in alcohol fermentations in Australian.89

For example, several kinetic parameters for a Z .mobilis fermentation were compared with Saccha- romyces carlsbergensis

10specially selected for its

sugar and alcohol tolerance.llBoth specific ethanol

productivity and specific glucose uptake rate are sev-

eral times greater for Z . mobilis. This result is mainly

due to lower levels of biomass formation and glucoseconsumption. The lower biomass produced wouldseem to be a consequence of the lower energy avail-

able for growth with Zymomonas than with yeasts—

the Enter-Doud oroff pathway produ cing only 1 moleof adenosine triphosphate (ATP) per mole of glucose,

compared to glycolysis with 2 moles ATP per mole

glucose. In none of the first three examples can etha-

nol be produced from pentose sugar.

The fourth approach utilizes a mixed culture of

Clostridia, which will utilize cellulose and hemicellu-

lose, hexoses, and pentoses for ethanol production.

The appl icat ion o f gene t ics for

improv ing mic rob ia l s t ra ins

In the previous sections, the process steps have

been identified that are particularly sensitive to thequality of the microbial strains. The following are im-

provements of microbia l charac te r i s t i cs tha t a re

either now possible or might be so in the future and

that will have an impact on the overall economics of

the process. The effect of new genetic techniques re-

quiring future research is similar for all micro-orga-

nisms in two w ays.

1. Manipulations could be attempted today with

less effort and greater chance of success if tools

like cell fusion and recombinant DNA (rDNA)

techniques were avai lable for al l of the mi-

crobes of interest.

——T;ibbs, et al., op. cit.‘K. J. Lee, D. E. Tribe, and P. L. Rogers, “Ethanol Production by Zymo-

monas mobilis in Continuous Culture at High Glucose Concentrations,” Bio-

technology Lett. 421-426, 1979.

9P. L. Rogers, K. J. ke , and D. E. Tribe, Biotechnoi. Lett. 1:165-170, 1979.

IOlbid.

I ID. Rose, Proc. Bichem. 11 [2), 1976, pp. IO-12.

2. Manipulations require further knowledge in a

specific area or the development of an entirely

new genetic system in ethanol producing mi-crobes—e.g., there is no genetic system for the

thermophilic anaerobic bacteria. Knowledge on

how to genetically alter ethanol tolerance of

both bacteria and yeast is lacking.

The economics of the fermentation of a substrateinto alcohol is primarily controlled by three factors:

1.

2.

3.

Ethanol yield.—The amount of product pro-

duced per unit of substrate determines the ma-

jor raw materials cost of the fermentation. Final ethanol concentration.—The cost of separat-ing the ethanol from the fermentation broth is a

funct ion of the ethanol concentrat ion in that

broth.

Productivity. —The amount of ethanol produced

per liter of fermenter volume per hour deter-mines the capital cost of the fermentation step,

once the type of fermenter and the annual out-

put have been chosen. Productivity is not inde-

pendent of the final ethanol concentration, andso an optimum compromise between these vari-

ables must be chosen.

T h e imp ac t o f ge ne t i cs on e tha no l y i eld

Most microbes that are chosen for making ethanol

already produce nearly the theoret ical maximum

yield. In these cases little improvement can be made.

The yield may be lower when the microbe hasbeen chosen for its other technical advantages such

as ability to degrade cellulose. Lower yield of a

microbial end product, like ethanol, can result from

the diversion of substrate to cell mass or to an alter-

native product. Both of these faults can be readily at-tacked. A number of cell changes (e.g., leaky mem-

branes) can cause the microbe to waste energy, re-

quiring it to metabolize more substrate into alcohol

to make the same cel l mass. Where the thermo-

dynamics and redox balance of the fermentat ion

allow, unwanted waste products can be eliminated

by mutation of the relevant pathways. Only limited

work has been done on this type of research with in-

dustrially significant bacteria.

T he impac t o f ge ne t i c s on f ina l e thano l

c o n c e n t r a t i o n

This is amenable to genetic manipulation, both em-

pirical and planned. An improvement in ethanol tol-

erance decreased both separation costs and ferment-

er capital cost (through increased productivity).

When traditional distillation is used, the effect on




complex molecules, ready-made, such as amino acidsand vitamins.

The more cheaply these nutrient needs can beprovided, the better. Whenever an organism can begiven genes from another source by applied biotech-nology techniques, there is a possibility that complexnu trient requ irements can b e obviated. However,

this requires that all the genes in a given pathway belocated in the source and be m ade to function in thenew microbes. The feasibility of this is un certain, butsolutions would decrease the cost of producing etha-nol with yeast as well as clostridia.

q Stain stability. —Many of the suggested ethanolprocesses propose to employ continuous culture.Although this offers several advantages over batchculture, it is somewhat vulnerable to deleterious mu-tations of the microbe u sed, particularly if the m i-crobe has been extensively altered in ways that makeit less competitive.

These deleterious genetic changes are almost en-tirely catalysed by biological systems in the microbe.

Alteration of these systems, so that the frequency of unwanted genetic changes is decreased, could great-ly extend the period of op eration th at is possiblebefore having to shut down and restart the fermen-tation. So far, this is a p ossibility only in microbesthat have a highly developed genetics. It may be thatstrain stabilization of this sort would not be possiblein other microbes until their genetics are highly de-veloped.

It is also possible to design strategies using currentstrain d evelopment techniq ues that might lead togenetically stable strains, but these are unproven.

q Cell separations. —Many fermentation schemesincorporate cell recycle to boost productivity. Thisrequires that cells be separated from effluent broth.Others need to separate cells from other residue as abyprod uct. In addition, some of the low-energy alter-natives to d istillation, such as adsorp tion, could re-quire separation of the cells from th e broth p rior toethanol recovery.

In these cases, microbes that can be made to floc-culate and red isperse, or that can be mad e to rever-sibly change their morphology would allow cheapgravity separations (settling or flotation).

q Operating conditions.—An increase in the

temperature an organism wil l tolerate is advanta-

geous for heat removal and in situ ethanol removal

schemes. The feasibility of accomplishing this is

uncertain.The extreme of productivity improvement via cell

recycle is an immobilized cell reactor, It is con-

ceivable that cel ls could be made less prone to

degradation under the conditions of immobilization,

by modifying sensitive components and degradation

systems, and by adding protective systems. This isnot at all a near-term possibility.

q Range and efficiency of substrate utilization.-Asingle-step conver sion of a substrate to ethanol ishighly desirable. This often requires that the ethanolfermenting organism possess a degradation capabili-ty it does not have.

As an example, consider ligno-cellulose. It consistsof hexosans, pentosans, and lignin, All of these com-ponents should be used. Assume that one cellulase-producing candidate does not use pentoses, while arelated noncellulase producing organism d oes, this isexactly the situation w ith clostridia. If the secondorganism can be given the cellulase genes of the first,a microbe better-suited to direct conversion could becreated. The pace at which such a manipulationcould be developed cannot be predicted with con-fidence, although this is not necessarily a long-termprospect.

Another obvious area that m erits attention is theenhancement of cellulase activity. Classical genetic

manipulations, employing mutation and selection orscreening, should result in m icro-organisms betterequipp ed to d egrade cellulose. E.g, it should be possi-ble to isolate strains that are deregulated in cellulaseproduction (hyperproducers) as well as those inwh ich the cellulase is not subject to p rodu ct inhibi-tion. In ad dition, it is tempting to think about thepossibilities of amplifying cellulase genes by meansof DNA technology and cloning. However, this latterapproach must aw ait further un derstanding of thebiochemistry and genetics of the cellulase system aswell as the development of the app ropriate geneticsystems in cellulolytic micro-organisms,

U t i li z a t i on o ff e r m e n t a t i o n b y p r od u c t s

Presently for each gallon of ethanol produced, ap-proximately 14 liters of stillage is formed.

17If ethanol

is mixed with gasoline to make gasohol (10 percentethanol), the total stillage produced annually in theUnited States would b e in the billions of liters. Surelya problem of this magnitude deserves serious atten-tion. The utilization of stillage or fermentation by-products could be greatly improved by geneticmeans in several ways, In actuality, only a rationallong-range genetic approach can increase the valueof such a fermentation byproduct. Value can be in-creased in two main ways. The first is to increase thenutritive value of the fermentation byprodu ct fol-lowed by developing economical processing technol-

i ~w . E. Tyner, “T he Pot en t j a l of O b t a i n i n g Energy F r o m A g r i c u l t u r e , ~W-

posium on Biotechnology The Energy Production and Conservation, Gatlin-

berg, Term., 1979.




ogies that stabilize and preserve nutritive value. The

second approach is to increase the functionality of

the byproducts so that more useful products can be

developed.For this one can envisage clever and novel ways to

utilize mutants to increase the value in a mannersimilar to those described.

18 l9 20Ethanol production

is not compatible with producing a valuable byprod-

uct. E.g., a filamentous yeast may be useful for directtexturization or fortification of an animal food but

production of ethanol may not be suitable with such

an organism. A possible solution to this type of con-flict involves the development and engineering of

two-stage fermentation processes. In the first stage,

ethanol producing organisms are propagated under

optimal economic conditions for ethanol production.After the production phase is over, the organisms

are then t ransferred to a second-s tage reac tor ,

where desirable phenotypic properties are then ex-pressed. Signals for expression of phenotypic prop-

erties can be extrinsic environmental parameters,

such as temperature, or levels of oxygen or carbon

dioxide, or intrinsic parameters, such as specific

nutrient requirements.

Thus the large-scale utilization of fermentation

byproducts as feed or other materials wil l then

become more valuable when genetic engineering can

decrease process ing cos t s and increase product

laA. J, Sinskey, J. Boudrant, C. Lee, J. DeAngelo, Y. Miyasaka, (;. Rha, and S.

R. Tannenbaum, “Applications of Temperature-Sensitive Mutants for Single-

Cell Protein Production,” in Proceedings of U.S./U.S.S.R. Conference on Mech-

anisms and Kinetics of Uptake and Utilization o f substrates in Processes for

the Production of Substances by Microbiological Means, Moscow-Pushchino,

p. 362, June 4-11, 1977. PB. 283-330-T.19J, Boudrant, J. DeAngelo, A. J. Sinskey, and S. R. Tannenbaum, “process

Characteristics of Cell Lysis Mutants of Saccharomyces cerviciae,” Biotech.

Bioeng. 21:659, 1979.ZOY, Mivasaka, A. J. Sinskey, J. DeAngelo, and C. Rha , “Characterization of

a Morph ological Mutant of saccharomyces cervisiae for Single-Cell Protein

Production,” ./. Food Science 45:558 ;563, 1980.

quality. Most of these types of studies remain to bedone. However, the potential for innovative applica-tions is great, but such applications may not result

because of the current lack of any Government agen-

cy that has a sound program for funding biotech-

nology research.

R e co m m en d a t i o n s a n d a r e a s i n w h i ch

app l i e d ge ne t i c s shou ld hav e an impac t

There has been little published research done inthe United States on the genetic improvement of ethanol production processes with bacteria such asZymomonas and clostridia, and only limited studieswith yeast. In light of p revious d iscussion, the follow-ing points have been identified as being the most im-portant and relevant in the app lication of geneticsfor improving ethanol-producing processes:

q

q

q

q

q

q

q

q

q

improvements on ethanol yield;increased ethan ol tolerance to achieve high er

final ethanol concentrations in the fermentationbroth;increased rates of ethanol production;elimination of unw anted produ cts of anaerobiccatabolism, that is, direction of catabolismtowards ethanol;enhanced cellulolytic and/or saccharolytic capa-bi li ties to improve rates of conversion of cellulose and/or starch to fermentable sugars;incorporation of pentose catabolic capabilitiesinto ethanol producers;development of strains capable of hydrolyzingcellulose and starch as well as of producingethanol from pentoses and hexoses;

improved temperature stability of micro-orga-nisms and/or their enzymes; andimproved harvesting properties of cellular bio-mass produced during fermentation.



Appendix II-A

A Case S tudy o f Whea t

Wheat is a major food staple in the diet of a large

percentage of the world’s population. Wheat grain in

the United States is used almost exclusively for

human consumption, although temporary localized

oversupply may result in some wheat feeding to live-

stock.

Attempts to improve wheat plant populations by

selection began several thousand years ago. The de-

sirable at t ributes selected included the abi l i ty to

withstand severe environmental st resses such as

heat, cold, and drought and the stability of the seed

head (which tends to disarticulate in wild forms).

Wheat seeds moved from country to countryalong with explorers and colonists. New varietiesplayed major roles in the establishment of many

productive wheat cultures—e.g., the Mennonite set-

tlers introduced hard red winter (Turkey Red) wheat

into the Kansas area from Russia in the late 19th

century. And two private breeders—E. G. Clark of Sedgenick, Kans., and Danne of Elreno, Okla.–de-

veloped varieties that set new levels of productivity

and straw strength in hard winter wheats which

were sought by millers for their excellent flour

recovery.

Breeding programs expanded during the first half

of the 20th century. At first, the U.S. Department of

Agricul ture (USDA) played a lead role; hut the

emergence of the Land Grant System and the estab-l ishment of the State experiment stat ion concept

prompted individual States to launch breeding pro-

grams designed to address the particular productionproblems faced by farmers within their respective

boundaries.

As the State experiment stations began to assume

more responsibility, USDA programs and personnel

began to concentrate in central locations to assemble

the optimal number of personnel for the greatest in-

teraction and productive output. If the present trend

continues, there will be virtually no USDA scientists

engaged in actual wheat breeding. Instead they will

have assumed the roles of basic researchers and re-

gional coordinators supplying information to the

public and private breeders.

Disease and insect resistance have been the pri-

mary breeding goals of many programs. The dramat-

ic losses associated with severe pest problems have

focused the attention of producers, researchers, andlegislators on these areas of need. Other traditional

breeding object ives have included improved use

properties, tolerance to environmental stresses such

as cold, wheat, wind dessication, and excessive mois-

ture, and inherent yield capacity in the absence of

significant production limitations.

The qual i ty of wheat’s end products has been

improved significantly through breeding. Varieties

have been tailored to meet the demands of various

industries. The bread bakeries needed a higher pro-

tein and more gluten strength to make a l ighter,

larger loaf, while the cookie producer needed a low-

protein flour with desirable dough-spreading prop-

erties.

W h e a t p r o d u c t i v i t y a n d m a n a g e m e n t

The pattern of wheat productivity (yield per acre)

in developed countries is remarkably similar. Whenyields are plotted over the centuries, there is a long

period of barely perceptible increases in yield, from

the time of first records of production to the end of

the first third of this century (the period of 1925-35).

Since around 1935, yield has increased sharply. Re-

cent data suggest that yield increases may be leveling

off. Why increases have been so substant ial after

generations of little success, is a complex question in-

volving genetic resources, economic development,

social interaction, and adoption of mechanical and

biological inn ovations.

Until recently, the U.S. commercial seed compa-nies, with one or two exceptions, have not been in-

terested in wheat breeding programs as a profitmak-ing venture. Since wheat has a perfect flower and

can fertilize itself, the farmer can purchase seed of a

new variety and reproduce i t from generat ion to

generation. However, the discovery of cytoplasmic

male sterility and nuclear restorer genes has stim-

ulated industry interest in the possibility of devel-oping hybrid wheat. The farmer would purchase the

hybrid seed each year; the inbred lines used to make

the hybrid would be the exclusive property of the

originat ing company. Although progress has been

good, problems exist with the sterility and restorer

systems, the ability to produce adequate amounts of

hybrid seed, and the identification of economic levels

of hybrid vigor. The next 5 years should reveal the

potential for success in hybrid wheat.

Several milestones of progress have been set inwheat. Yield has risen dramatically. Genetic protec-

tion against pests and other hazards has been a m a- jor contributor to increased yields. In addition, re-

cent advances using semidwarf genes have been as-

304



Appendix II-A—A Case Study of Wheat . 305

sociated with significant yield improvement . The

shorter, stiffer stems of the semidwarf plants allow

maximization of resources without yield reductions.

Improvement in the inherent yield components of stems per uni t area, kernels per stem, and kernel

weight has also contributed extensively to yield im-

provement.

The use of applied genetics in wheat improvement

occurs in close harmony with total wheat manage-

ment systems. The farmer must integrate a huge as-

sortment of alternatives in each decision—e.g., an in-

dividual producer may be deciding on a nitrogenprogram. I f the fa rm i s i r r iga ted, the producer

selects ni t rogen amounts and applicat ion t iming

based on soil tests, intended crop and variety, the

end use of that crop, and watering schedules. [f the

farm is rainfed, the producer takes into account soil

tests, crop considerations, and rainfall probabilities.

In both cases product prices at the time of sale

must be predicted since they govern potential gross

return, which in turn affects the costs of maintaining

a profit margin. Genetic interaction in this system isintricate. The farmer must first select the variety

most likely to produce at the maximum economic

level. For irrigated land, it may be a short high-

yielding semidwarf either for the cookie trade or theexport market. The farmer knows that part of the

value of his product is dependent on low protein.

However, inappropriately high levels of ni t rogen,

which greatly improve yield, will also raise the pro-

tein of the crop beyond acceptable levels. If the ex-

port market is strong and the total U.S. supply re-

duced, the higher protein may be of little economic

consequence.In the case of the dryland farmer, the variety

selected may be taller with lower yield potential but

with much better levels of adaptation and tolerance

to adverse environments. It may be designed for the

bread industry or the export market . Part of the

value is related to high-protein content. Since mois-

ture conservation and use is critical, nitrogen ap-

plications and amounts must be selected so that the

plants do not waste their moisture reserve. However,nitrogen applied too late may not receive enough

rain to penetrate the soil and become available to the

plants. If the plants “burn up” because of unwise

water use early in the season, the seeds will be high

in protein but low in yield. If inadequate nitrogen is

available, the crop will generally be low in protein.

The abbreviated protein story is but one of manyexamples of farm management interaction with ap-

plied genetics in wheat production. Recent changesin energy price and availability, environmental re-

straints, marketing structures, and technology devel-

opment are producing a new array of complex prob-

lems.

Gene t i c v u lne rab i l i t y in w he a t

Genetic vulnerability is defined as a high degree of

genetic uniformity in a crop grown over a wide acre-age. Wheat, which is produced on about 62 million

acres annually in the United States, has a relativelyhigh level of uniformity and genetic vulnuerability.

In 1974,102 hard red winter wheat varieties were

grown on 36.6 million acres, with four varieties oc-

cupying 40 percent of the acreage. Hard red spring

wheat varieties totaled 80 percent on 14.7 million

acres, with three varieties occupying 52 percent of

the acreage. Similar situations occurred with other

classes of wheat. Plant pests, including diseases and

insects, have periodically caused moderate to servere

wheat crop losses in years favorable to the develop-

ment of strains capable of attacking current forms of

resistance.

Incorporating genetic resistance to pests has tradi-

tionally been the responsibility of public breeders.

Wheat is a self-fertilized plant that can be faithfully

reproduced from generation to generation. Private

industry has been reluctant to invest R&D money in

improvements since the farmer, following the initial

seed purchase, can reproduce the crop without re-

turning to the seed company. Thus, public breeders

have been the main source of new varieties and have

had the responsibi l i ty of del ivering genet ic im-

provements to the producer. Wheat breeding pro-

grams are generally designed to respond to State pro-

duction needs. Goals and objectives are established

by technical advisory groups that include breeders

and sc ient is t s, growers , use indus t ry representa -

tives, and extension workers.

Genetic variability is available to the breeder from

naturally occurring sources and artificially induced

mutations. Naturally occurring variability has been

collected from native plant populations throughoutthe world and is maintained in the World Wheat C ol-

lection by the Science and Education Administrationof USDA located in Beltsville, Md. Currently, about

37,000 accessions are contained in the collection.

Breeders use the collection as a reservoir from which

to draw exotic genes needed to improve the value of

their breeding programs. In addition to variability

within wheat varieties, the breeders can use special

genetic techniques to draw valuable genes from re-

lated species such as rye and various forage grasses.

This approach, while time-consuming and costly, has

been used in a number of variety development pro-

grams. Mutations induced by artificial means have



306 q lmpacts of Applied Genetics-Micro-Organisms, Plants, and Animals

not been used extensively by the breeders, since ly evaluated, characterized, and documen ted, forc-

desired mutations without detrimental effects are ing breeders to spend time and resources carrying

very difficul t to obtain. Enough natural genet ic out their own evaluation work. The committee has

variability seems to exist to satisfy needs in the proposed a standard set of descriptors for all acces-

foreseeable future. sions in the collection, as well as an information

The National Wheat Improvement Committee has management system to efficiently bring the informa-

stated that the World Wheat Collection is inadequate- tion to the breeders.



Appendix II-C

A ni ma l F e r t i l i z a t i on Techno l og i e s

Spe rm s torage

DEFINITION

The freezing of semen to – 196° C, storage for anindefinite time, followed by thawing and successful

insemination.

STATE OF THE ART

Conception rates at first insemination with frozensperm average between 30 to 65 percent for mostspecies. This technology is not a key to the success of artificial insemination (AI), but because of the con-venience it is now an essential ingredient. Current

operational procedures ar e adequate for the dairy in-

dustry.

ADVANTAGES1. Greater use of selected bulls as AI studs.

2. Elimination of the n eed to maintain expensive and

dangerous bu lls on dairy farms.

3. Sperm can be tested for disease and treated for

venereally transmitted diseases.

4. Ease of transport and therefore of increasing po-

tential offspring.

FUTURE

Little change is anticipated in semen processing.Freeze-dried semen is unlikely to be successfulenough to use. Sperm banking can be expected to in-crease, especially on AI studs. Banking provides

cheap storage while bulls (slaughtered) are beingprogeny tested, and insurance against loss of bullsthrough natural causes. For preservation of semenfrom bulls of less populous breeds, banking can becompleted in about a

-year, after

be slaughtered.

Ar t i f i c ia l i n sem ina t ion

DEFINITION

which the-bull can

Manual placing of sperm into the uterus.

STATE OF THE ART

Highly developed for most species. Representativeuse rates in the United States are: dairy cattle, 60 per-

cent; beef cattle, 5 percent; turkeys, 100 percent.

The major limitation to the use of AI is the low na-

t iona l average concept ion ra te a t f i rs t se rvice ,around 50 percent. The success or failure of AI is

determined by a mult ipl ici ty of factors including

estrus detection, quality of semen, timing of in-semination, and semen handling.

DISADVANTAGES1. With increased herd size, estrus detection has

become a major problem.2. Inexperienced dairymen are buying semen and in-

seminating their own cows, resulting in loweredfertility and n o feedback on semen fertility.

ADVANTAGES

1. Widespread use of genetically superior sires.2. Services of proven sires at a lower cost.3. Elimination of cost and d anger of keepin g bu lls on

the farm.4. Control of certain diseases.S. Use of other breeding techniques including cross-

breeding.6. Continued u se of valuable sire after his d eath.

FUTURE

Greater use of AI in beef cattle will depend on theavailability of successful and inexpensive estrus syn-chronization technology, on relaxed restrictions of the various breed associations, and on accurate prog-eny records,

Es trus synchronizat ion

DEFINITION

Estrus (“heat”), is the period during which thefemale will allow the male to mate her. This sexualbehavior is subtle and varies widely among in divid-uals. Thus the synchronization of estrus in a herd,using various drug treatments, greatly enhances AIand other reproduction programs.

STATE OF THE ART

Effective method s for synchronization of estru speriods for large numbers of animals have beenavailable for more than two decades, and several ap-proaches are now available which result in normalfertility. Several schemes involve use of prosta-glandin F

2(PGF

2) for the cow and ewe. However, FDA

approves usage only for controlled breeding in beef

cows and heifers, nonlactating dairy heifers, and inmares.

ADVANTAGES

1. Time a heifer’s entry into a milking stream.2. Increase productivity by breeding heifers earlier

in life.

309

76-565 0 - 81 - 21




3.

4.

toin

Ability to breed large numbers of cattle over ashorter calving interval.Increase use of AI, especially in beef cattle, sheep,and swine.

FUTURE

Estrus cycle regulation should allow selected sires

be more widely used to improve important traitsbeef cattle. It shou ld also gain wid espread an d

rapid acceptance among d airymen as well.-

Supe rov u la t ion

DEFINITION

Superovulation is th e hormonal stimulation of multiple ovarian follicles resulting in release fromthe ovary of a larger n um ber of oocytes (ova) thannormal.

STATE OF THE ART

Superovulation with implantation into surrogatemothers increases the number of offspring, usuallyfrom highly selected dams. Adequate procedures arepresently available for superovulation of laboratoryand domestic animal species, except the horse. Thedrugs used to induce superovulation are the go-nadotropins, pregnant mare’s serum gonadotropin(PMSG) and follicle stimulating hormone (FSH), insome instances followed by other treatments to stim-ulate ovum maturation and ovulation. Superovulatedova result in normal offspring with the same successrates as achieved w ith n ormally ovulated ova.

DISADVANTAGES

1. Greatest drawback is that degree of success can-not be p redicted for an individu al animal.2. Batches of hormones for ovulation treatment vary

widely in q uali ty.3. PMSG is scarce, and has been declared a drug by

the Food and Drug Administration (FDA). Thus,most use of PMSG is now illegal.

4. There is insu fficient d ata to jud ge the effect of repeated superovulation.

FUTURE

Methods for superovulation will imp rove consist-ency of results. Add itional und erstandin g of basicphysiological mechanisms will facilitate such efforts.New work in superovulatory technology involves ac-tive immun ization against adrostenedione (a hor-mone involved in regulation of follicular develop-ment). This treatment prevents atresia and reliablyincreases the frequency of multiple ovulations. Thetechnology has definite commercial potential for cat-tle husbandry and limited potential for sheep hus-

bandry, and much current effort is directed towardsdeveloping and testing a commercial procedure.

E m b r y o r e c ov e r y

DEFINITION

The collection of the fertilized ova from theoviducts or uteri . Collection of embryos is anecessary step for emb ryo transfer or storage, andfor many experiments in reproductive biology. Bothsurgical and nonsurgical methods are used.

STATE OF THE ART

Surgical. -Method s are available for recovering40 to 80 percent of ovulations from cattle, sheep,goats, swine, and horses. The development of adhe-sions and scar tissue following surgery limits thesetechniques. Surgical recovery is the only method forsheep, goats, and pigs. It is presently practicedalmost exclusively when a susp ected p athology of theoviducts renders an individual subfertile, or whenembryos must b e recovered b efore the ind ividualreaches puberty.

N o n s u r g i c a l . - Non-surgical embryo recoverytechniques are preferred for the cow and horse.Fifty to eighty percent of cow ovulations can berecovered, and 40 to 90 percent of the operations onhorses to recover the single ovulation are successful.

ADVANTAGES

1. Nonsurgical embryo transfer can be performed anunlimited number of times.

2. Requirements for equipm ent, personnel, and timeare low in nonsu rgical recovery. This is esp eciallyimportant in milk cattle: since the non surgical pro-cedure is p erformed on the farm, milk p roductionis not interrupted.

3. A single embryo can be obtained between super-ovulation treatments.

4. Embryos can be obtained from a young heiferbefore it reaches puberty.

5. The technology is especially important for re-search, e.g., in efforts to produce identical twins,embryo b iopsies for sex determination, etc.

FUTURE

Methods of collecting embryos have not changedapp reciably sin ce about 1976, nor are significant ad-vances predicted for the future.

Embry o t rans fe r

DEFINITION

Implantation of an emb ryo into the oviduct oruterus.



Appendix II-C—Anim81 Fertilization Technologies q 311

STATE OF THE ART

Surgical.—Pregnancy rates of 50 to 75 percentare achievable in cows, sheep, goats, pigs, andhorses. Surgical transfer is the only practical methodin sheep, goats, and pigs, and is the predominantmethod for cows and horses. A nu mb er of factors

determin e the su ccess of surgical transfer: age andqu ality of embryos, site of transfer, degree of syn-chrony between estrous cycles of the donor and re-cipients, number of embryos transferred, in vitroculture conditions, skill of personnel, and manage-ment techniques. The 50- to 60-percent success ratein cattle compares with AI success rates at first serv-ice. (Pregnan cy rates should not b e confused w ithsurvival rates, which m ay be much lower.)

Nonsurgical . —This method is an adaptation of

AI. Reported success rates are much lower than

those with surgical transfer. Nonsurgical transfer is

not u sed in sh eep, goats, or pigs.

1.

2.

3.

4.

5.

6.

7.

1.

2.

ADVANTAGES

Obtaining offspring from females unable to sup-port pregnancy,Obtaining more offspring from valuable females.With a homozygous donor, undesirable recessivetraits among animals u sed for AI can b e rapidlydetected.Introducing new genes into specific pathogen-freeswine herds.Coupled with short- or long-term embryo storage,transportation of animals as embryos.Increasing the population base of rare or endan-gered breeds of animals by use of closely relatedbreeds for recipients.Separation of embryonic and maternal influencesin research.

DISADVANTAGES

Personnel requirements in surgical transfer ac-count for a large share of high costs and thus limitapplicability in animal agriculture.Provision of suitable recipients is the greatestsingle cost in embryo transfer.

FUTURE

Surgical transfers will remain the method of choice for sheep, goats, and pigs in the foreseeablefuture. For cows and horses, however, nonsu rgicalmethods will be increasingly used rather than sur-

gical techniques (and this will be apparent) withinthe n ext year or two. It is likely th at half of th e com-mercial transfer pregnancies in cattle in North Amer-ica in 1980 will be done nonsurgically, even if suc-cess rates are only 60 to 80 percent of those obtain-able with surgical transfer. Among future appli-cations, a role for embryo transfer can be predicted

in p rogeny testing of females, obtaining twins in b eef cows, obtaining p rogeny from prepub ertal females,and in combination with in vitro fertilization and avariety of manipulative treatments (e.g., productionof identical twins, selfing, genetic engineering, etc.)

Embry o s to rage

DEFINITION

Maintenance of embryos for several hours or days

(short-term) or for an indefinite length of time (freez-ing).

STATE OF THE ART

S hor t - t e rm . -T he r equ i r em ent fo r em bryosfrom farm animal species has not been defined,although adequ ate culture systems for the short in-terval between recovery and transfer have beendeveloped by trial and error. Whereas the importantparameters of culture systems have been identified

(e.g., temp erature, pH , etc.), optimal conditions havenot been determined. Cow embryos may be storedfor three days in the ligated oviduct of the rabb it.

Long-term (freezing).-No completely adequateprotocol exists for freezing emb ryos of farm sp ecies.One-third to two-thirds of embryos are killed usingpresen t meth ods. Pregnancy rates of 32 to 50 per-cent for cattle, sheep, and goats have been reportedafter freezing. No successful freezing of sw ine orhorse embryos followed by development to term hasbeen rep orted. Despite disadvan tages (one-half of embryos are often killed) advantages are such that insome’ situations embryo freezing, and embryo sell-ing, are already profitable.

ADVANTAGES

1. Amplification of advantages of embryo transfer.2. Elimin ation of req uiremen ts for large recipien t

herds when embryo transfer is being used.3. Reduction of costs in animal transport.4. Control of genetic drift in animals over prolonged

time intervals.

FUTURE

Anticipated development of embryo culture tech-nology would be of significance in efforts toward invitro maturation of gametes, in vitro fertilization, sexdetermination, cloning, and genetic engineering, all

of which involve prolonged manipulation of gametesand embryos outside of the reproductive tract.

As freezing rates improve, nearly all embryos re-covered from cattle in North America will be frozen.Probably as man y as half of the embryos will bedeep-frozen for 2 to 3 years. It is unlikely that suc-cess rates will ever approach 90 percent of those




without freezing. However, 70- to 80-percent successrates may be attainable within several years. It ap-pears that embryos can be stored indefinitely withlittle deterioration.

Sex s e l e c t i o n

DEFINITION

Tests to determine the sex of the unborn or deter-mination of sex at fertilization by separating x- bear-ing from y-bearing sperm.

STATE OF THE ART

Sexing of embryos.—Through karyotypingnearly two-thirds of em bryos can b e sexed. Tech-niques using identification of the condensed X chro-mosomes are unreliable. A third method, identifica-tion of sex-specific gene products, is under develop-ment.

Sexing of sperm. —A 100-percent meth od has

not been achieved in any mammalian species; and nostandard protocol for farm species exists.

FUTURE

Before this technology can be applied commercial-ly, it must be simple, fast, inexpensive, reliable, andnonharmfu l for embryos. Such techniq ues could u n-doubtedly be developed. There would be numerousmedical and experimental applications.

There is much interest in research in this areabecause of its use in u nderstanding male fertilitywith AI in humans, and in enhancing sperm survivalafter frozen storage.

T w i n n i n gDEFINITION

Artificial production of twins, either using embryotransfer or hormone treatments,

STATE OF THE ART

Currently, embryo tran sfer is the most effectivemethod for inducing twin pregnancies in cattle,resulting in pregnan cy rates of betw een 67 to 91 per-cent, of which 27 to 75 percent deliver twins. Othermethods include transferring one embryo into a cowwhich has been artificially inseminated, and hor-monal induction of twinning, which is a modification

of superovulation. This latter method is not reliable.ADVANTAGE

The advantage of twinn ing in nonlitter-bearingspecies is the improved feed conversion ratio of pro-ducing the extra offspring.

DISADVANTAGE

Th e major disadvantage of twinning is intensivemanagement necessary for periparturient complica-tions, unpredictable gestation periods, depressed lac-tation, etc.

FUTURETechnical feasibility for twinning, in conjunction

with embryo transfer, management adjustments,and selection for good recipients, can be predicted. Areliable procedure for twinning in sheep can also beexpected. The technology would most likely be firstused in Europe and Japan, where there are shortagesof calves to fatten for beef.

In v i t ro fer t i l i za t ion

DEFINITION

The union of egg and sperm outside the reproduc-

tive tract. For some species, the technology includessuccessful development of the embryo to gestationand birth.

STATE OF THE ART

In vitro fertilization has been accomplished inseveral laboratory animal species, including the rab-bit, mouse, rat, hamster, and guinea pig an d n ineother mammalian nonlaboratory species, includingman , cat, dog, pig, sheep, and cow. However, norm aldevelopment following in vitro fertilization and em-bryo transfer has only been accomplished in the rab-bit, mouse, rat, and human. Consistent and repeat-able success with in vitro fertilization in farm specieshas not yet been accomplished.

Non e of the cases of reported su ccess of in vitrofertilization, embryo transfer, and normal develop-ment in man is well documented.

Most of the in vitro fertilization work to date hasconcentrated on the development of a research toolso that the physiological and biochemical events infertilization and early development could be betterun derstood. More practical app lication of in vitrofertilization techniques would include:

1. a means for assessing the fertility of ovumand/or sperm;

2. a means to overcome female infertility with em-bryo transfer into a recipient animal; and

3. when coupled with ovum and/or embryo stor-age and transfer, a means to facilitate combina-tion of selected ova with selected sperm for pro-duction of individuals with predicted character-istics at an appropriate time.



Appendix /l-C—Animal Fertilization Technologies q 313

FUTURE

Rapid progress in research is anticipated andmany of the potential applications of in vitro fertiliza-tion to animal breeding should become practicalwith in th e next 10 to 20 years. With further d evelop-ment of in vitro fertilization methodology, along withstorage of unfertilized oocytes (gamete banking), fer-tilization of desired crosses should become possible.In the more distant future, genetic engineering andsperm sexing along with in vitro fertilization maybecome possible.

P a r t h e n o g e n e s i s

DEFINITION

Th e initiation of development in the absense of sperm.

STATE OF THE ART

Parthenogenesis has not been satisfactorily dem-

onstrated or described for mammalian species. Thebest available information leads to the conclusionthat maintenance of parthenogenetic development toproduce normal offspring in mammals approximatesimpossibility.

C lon in g: p rodu c t ion o f i de n t i c a l tw in s

DEFINITION

The production, using a variety of methods, of genetically identical individuals.

STATE OF THE ART

There are several ways to obtain genetically iden-

tical livestock. The natural way is identical twins,althou gh th ese are rare in sp ecies other than cattleand primates. Both natural and laboratory methodsdepend on the fact that the blastomeres of early em-bryos are totip otent (i.e., each cell can d evelop in to acomplete individ ual if separated from the others.)For practical purposes, highly inbred lines of somemammals are already considered genetically iden-tical; F1 crosses of these lines are also considered

genet ical ly ident ical and do not suffer from the

depressive effect of inbreeding.

ADVANTAGE

An advantage of identical twins is the experimen-

tal control provided by on e animal through whichtwo sets of environmen tal cond itions can be com-pared for effects on certain en d p oints, e.g., native v.surrogate uterine environments for gestational de-velopment, nutrition on milk production, etc.

C lon ing : nuc l e ar t r ans p lan ta t i on

DEFINITION

The production of genetically identical mammalsby inserting the nucleus of one cell into another,before or after destroying the original genetic com-plemen t. These occur by sep aration of embryos orparts of embryos early in development but well afterfertilization has occurred.

STATE OF THE ART

Experimentalists have found in certain amphibiathat transplantation of a nucleus from a body cell of an embryonic (tadpole) stage into a zygote followingdestruction or removal of the normal nucleus canlead to d evelopment of a sexually mature frog.

FUTURE

The ideal technique for making genetic copies of any given outstanding adult mammal would involveinserting somatic (body) cell nuclei into ova, which

may take years of work to p erfect if indeed it is possi-ble, There is some eviden ce that adult b ody cells areirreversibly differentiated.

How identical will clones be? They can be ex-pected to be fairly similar in appearance. They wouldbe less similar than identical twins, however, whichshare ooplasm and u terine and neonatal environ-ments. Furthermore, certain components are inher-ited exclusively from th e moth er, e.g., the m itochon-drial genome and perhaps th e genome of centrioles.The random inactivation of one or the other of the Xchromosomes may also limit similarities. Other dif-ferences among clones would result from the pre-natal environment: in litter-bearing species even

uterine position can affect offspring. In single-bear-ing species the maternal effect may be pronounced.Environmental differences in later life may greatlyaffect certain traits, even if those traits have a stronggenetic component.

Serious technical barriers must be overcomebefore realistic speculation of possible advantages inanimal production can be foreseen.

cell f u s i o n

DEFINITION

The fu sion of two m ature sex cells or the fertiliza-

tion of one ovum with another. An analogous schemefor the male wou ld b e accomplished by m icrosurgi-cal removal of the female pronucleus and substitu-tion of nuclei from two sperm. Combining sex cellsfrom the same animal is called “selfing.”




STATE OF THE ART

Combination of ova has led to early developmentto the blastocyst stage in the m ouse bu t no furtherdevelopment following transfer has been reported.Initial success in experimentation with manipulationof pronuclei has been reported.

FUTURECell fusion technology may someday prove useful

for getting genetic material from a somatic cell into afertilized l-cell embryo for the purpose of cloning. Inconjunction with tissue culture technology the tech-nology would have a role in gene mapp ing of chro-mosomes for the cow and perhaps other species.

Combining ova of the same animal, selfing,would rapidly result in p ure genetic (inbred) lines foruse as breeding stocks. The technique would alsolead to rapid identification of undesirable recessivetraits which could be eliminated from the species.

C h i m e r a s

DEFINITION

A chimera is an animal comprised of cell linesfrom a variety of sources. They can be form ed b yfusing two or more early embryos or by adding extracells to blastocysts.

STATE OF THE ART

Live chimeras between two species of mouse havebeen p roduced. Such young h ave four p arents in-stead of two; hexaparental chimeras have also beenproduced.

FUTURE

Practical app lications of chimera techn ology tolivestock are not obviou s at this stage of develop-ment. The main objective of this research is to pro-vide a genetic tool for better understanding of devel-opment, and maternal-fetal interactions.

Re c ombinan t DN A

DEFINITION

The introduction of foreign DNA into the germ-plasm.

STATE OF THE ART

The mechanics of changing the DNA molecules of farm animals directly have not yet been worked out.The plasmid methods used in bacteria may not be ap-plicable,

FUTURE

None of these techniques, no matter how great the

potential, will be of any use in animal breeding untilkn owledge of gen etics is greatly advan ced. Beforeone can alter genes, they must b e identified,

Prior to exploitation of recombinant DNA technol-ogy in anim al breedin g, it is necessary to identifygene loci on chromosomes, i.e., genetic mapp ing.Work toward this goal has only recently been initi-ated and rapid progress cannot be anticipated. Multi-variate genetic determinants of characteristics of economic importance are anticipated to be the rule.



Appendix III-A

H i s t o ry o f t he

R e c om b in a n t D NA D eb a t e

The history of the debate over the risks fromrDNA techniqu es and the Governm ent’s respon se

may be divided into four phases. * Phase I covered

the per iod f rom the f i rs t awareness of r i sks tohuman health from experiments involving recombi-

nant DNA (rDNA) in the summer of 1971 to the end

of the Conference at the Asilomar Center in Feb-

ruary 1975, which resulted in prototype guidelines

covering the research, Phase 11 covered the period

from Asilomar through the development by the Na-

tional Institutes of Health (NIH) of the Guidelines of

June 1976. In this period, the public first became

significantly involved in the debate and most, if not

all, of the policy issues were clearly framed. Phase

111, from mid-1976 through mid-1978, involved con-gressional consideration of the issues in an atmos-

phere that went from almost imminent passage of

legislation to the cessation of such efforts. Phase IVcovers the postlegislative period, when NIH and itsorganizational parent, the Department of Health,Education, and Welfare (HEW) (now the Departmentof Health and Human Services) undertook to developsatisfactory voluntary standards in areas over whichthey had no legal authority and to accommodategrowing pressure for public involvement, whileavoiding a full regulatory role.

Phase I began in the su mm er of 1971, wh en sev-eral scientists became concerned about the safety of

a proposed experiment to insert DNA from SV40virus, a monkey tumor virus that also transforms hu-man cells into tumor-like cells, into a type of bacterianaturally found in the human intestine. Aftermonths of discussion, the scientist who had pro-posed the experiment decided to defer it. Meanwhile,as rDNA techniques became more refined, debatesabout safety increased; at the June 1973 GordonResearch Conference, safety issues were discussed.The p articipan ts voted: to send a letter to the N a-tional Academy of Sciences (NAS) and the National In-

“For a detailed h istory through 1977, see footnote 1. For a his-tory and a discussion of the broader issues, see footnotes 2 and 3.

‘J. Swazey , J. Sorenson, and C. Wong, ‘(Risks and Benefits,

Rights and Responsibilities: A Historv of the Recombinant DNA Re-search Controversy,” Southern California Law Review SI:1019,

Septem ber 1978.‘C. Grobs t e i n , A Double Image of the Double Helig (San Fran-

cisco: W. H. Freeman Co., 1979).3D. Jackson, and S. Stich (eds. ), The Recombinant DNA Debate

(Englewood Cliffs, N . J.: Prentice-Hall, Inc., 1979).

stitute of Medicine>

requesting the appointment of committees to study potential hazards to laboratoryworkers and the pu blic; and by a n arrow majority4

to arrange for the letter to be published in the widelyread journal, Science, to alert the broad er scientificcommunity.

5

NAS appointed a committee of prominent scien-tists involved in rD NA research. In July 1974, thepanel asked for a temp orary worldwide moratoriumon certain types of experiments, and called for an in-ternational conference on potential biohazards of the research through a letter pu blished in Scienceand its British counterpart, Nature.

6 This letter also

requested the Director of NIH to consider estab-

lishing an advisory committee to develop an experi-mental p rogram to evaluate potential hazards andestablish guidelines for experimenters.

In response, the Director of NIH, after authoriza-tion by the Secretary of HEW, established the Recom-binant DNA Molecule Program Advisory Committee

(later renamed the Recombinant DNA Advisory Com-mittee, RAC) on October 7, 1974, along the lines sug-gested by the NAS Committee. The Committee’scharter described its purpose as:

7

The goal of the Comm ittee is to investigate the cur-rent state of knowledge and technology regardingDNA recombinant, their survival in nature, andtransferability to other organisms; to recommendprograms of research to assess the possibility of spread of specific DNA recombin ant and th e possiblehazards to pub lic health and to the environment; andto recomm end gu idelines on the b asis of the researchresults. This Committee is a technical committee, estab-lishedto look at a specific problem. (Emphasis added.)The internat ional conference cal led for by the

NAS Committee letter was held at the Asilomar Con-

ference Center, Pacific Grove, Calif. , in February

1975. The organizing committee made it clear that its

pu rpose was to focus on scientific issues rather thanto become involved in considering ethical and moralquestions. However, in one session the few lawyers

4Swazey, et al., op. cit., p. 1,023.

‘Letter from Maxine Singer and D ieter Soil to the NationalAcademy of Sciences (NAS ) and the National Institute of Medicine,reprinted in Science, vol. 181, 1973, p. 1114.

%etter from Pau l Berg, et al. to the editor, reprin ted in Science,

VO]. 185, 1974, p. 303.Th e charter of the Recombinan t DNA M olecule Program Ad-

visory Co mm ittee, Oct. 7, 1974.

315




invited confronted the scientists with some of thesequestions.’ The conference report concluded that al-though a moratorium should continue on some ex-perimen ts, most work in volving rDNA could con-tinue with ap propriate safeguards in the form of physical and biological containment,

In Phase II, th e debate widened to encompassbroader social and ethical issues, such as the re-lationship between scientific freedom of inquiry andthe protection of society’s interests, in whatevermanner those were defined. Such issues led natural-ly to questions about who makes the decisions andthe role of the pu blic in th at process. Finally, deci-sionmaking m echanisms w ere developed. Issuesraised and actions taken during this phase in manyrespects controlled the subsequent development of the Federal response to the debate, and createdproblems that continue to the present. At this stage,participation in the debate went beyond the scien-tific community.

Qu estions of ethics and p ub lic’ policy had beenraised earlier, but they now received much wider at-tention. On April 22, 1975, Sen. Edward M. Kennedy,Chairman of the Sub committee on Health of theSenate Committee on Labor and Public Welfare, helda half-day hearin g on science policy issues arisingfrom rDNA research. In May 1975, a 2-day con-ference on “Ethical and Scientific Issues Posed byHuman Uses of Molecular Genetics” was held underthe joint spon sorship of the N ew York Academy of Sciences and the In stitute of Society, Ethics, and th eLife Sciences. In addition to molecular biologists, par-ticipants included lawyers, sociologists, psychiatrists,and philosophers.

The issue of public participation arose as decision-making mechanisms were developed. RAC was orig-inally composed of 12 members from “the fields of molecular biology, virology, genetics and microbiol-Ogy.”

9Critics first noted the need for more expertise

in the fields of epidemiology and infectious diseases,since most molecular biologists were trained aschemists. * RAC’s membership was increased to 16and the range of expertise was widened to includethe fields of epidemiology, infectious diseases, andthe biology of enteric organisms, by amen dmen t tothe charter on April 25, 1975.

Since some members were conducting the re-search in question, critics claimed that a conflict of interests existed. They also noted that the Committee

%3wazey, et al., op . cit., p. 1,034.Whe charter of the Recombinant D NA Molecule Program Ad-

visory Co mm ittee, Oct. 7, 1974, op. cit.q One of the mem bers of the original RA C (Stanley Falkow) d id

have substantial expertise with ente r i c organisms and E. coli inparticular.

advised the Director of NIH, an agency whose mis-sion w as to foster biomed ical research, not to stop orotherwise regulate it. These issues were brought outin a petition to NIH signed by 48 biologists in August1975. Criticizing a proposed draft of the guidelines assetting substantially lower safety standards thanthose accepted at Asilomar, the petition argued forbroader representation on RAC from other fields of scientific expertise and from the public-at-large. RACitself had been sensitive to these limitations; in thesummer of 1975, an attempt was made to recruitnonscient is ts .

l0One nonscientist was added in

January 1976, and another was added in August1976.

In December 1975, RAC submitted revised draftguidelines to the Director of NIH, Dr. DonaldFredrickson, Although they were stricter than thosedrafted at Asilomar, some criticized th em as bein g“tailored to fit particular experiments that are al-ready on the d rawing boards.”11 The consensus of RAC, on the other han d, was that the gu idelines wereexcessively strict, but that it was necessary to beoverly cautiou s because of its limited expertise inpublic health.

l2In any event, Dr. Fredrickson ar-

ranged for public hearings on the proposed guide-lines at a 2-day m eeting in February 1976 of the Ad -visory Committee to the Director, a diverse group of scientists, physicians, lawyers, philosopher s) andothers. A similarly diverse group of scientists andpublic interest advocates were invited to attend.Some modifications to the Guidelines proposed byDr. Fredrickson as a result of that meeting wereadopted and others were rejected by RAC in April1976.

13

The final major issue arising d uring this periodconcerned NIH’s lack of authority to set conditionson research funded by other Federal agencies or bythe private sector. In a June 2, 1976, meeting be-tween Dr. Fredrickson and some 30 representativesof industry, including pharmaceutical and chemicalcompanies, it became clear that some rDNA researchwas being done; however, the representatives ap-peared hesitant to commit themselves to voluntarycompliance with the proposed guidelines.

l4The pri-

I O D r + E]izabeth Kutter, a former RAC member, persona] cot’n-

mun i c a t i on , Sep t. 11, 1980.1 I N , w a d e , ~ , Re ~ om b i n a n t DNA: N IH s e t s s t r i c t R u l e s t o ~unch

New Techn ology,”Science, vo l. 190, 1975, pp . 1175, 1179.

‘zKutter, op. cit.‘sIbid.l t subcommit tee on science, Research and Technology of the

House Committee on Science and Technology, Genetic Engineering

Human Genetics, and Cell Biology: DNA Recombinant Molecule Re-

search (Supp. Report 11) 94th Cong,, Zd sess,, 1976, p, 51.



Appendix III-A—History of the Recombinant DNA Debate q 317

mary reason was their concern over protection of proprietary information.

l5

Phase 11 culminated w ith the promulgation onJune 23, 1976, of the Guidelines for Research Involv-ing Recombinant DNA Molecules (“1976 Guidelines”)covering institutions and individuals receiving NIHfunds for this research.

Phase III was characterized by attempts to remedythe limited ap plicability of the Gu idelines. Soon aftertheir publication, Senators Kennedy and Javits sent aletter to President Ford, calling his attention to theGuidelines. They noted that any risk was not limitedto federally funded research, and urged him totake necessary steps to imp lement the G uidelinesthroughout the research community. In October1976, the Secretary of HEW, with the approval of thePresident, formed the Federal Interagency AdvisoryCommittee under the chairmanship of the Directorof NIH to determine the extent to which the G uide-lines could be applied to all research and to rec-ommend necessary executive or legislative actions toensure compliance.l6 In M arch 1977, the Committeeconcluded that existing Federal law would not per-mit the regulation of all rDNA research in the UnitedStates to the extent deemed necessary;17 it furtherrecommend ed new legislation, specifying the ele-ments of that legislation, is

Du ring 1977, several bills to deal with this an dother problems were introduced in Congress. Theyaddressed in different ways the issues of the extentof regulatory coverage, the mechanisms for regula-tion, and Federal preemption of State and local regu-lation. The major bills were those of Rep. PaulRogers, H.R. 7897 (and its substitute, H.R. 11192) andof Sen. Edw ard Kenn edy, S. 1217. *

While hearings were being held, three devel-opm ents occurred wh ich, by the en d of 1977, haddissipated mu ch of the im petus for legislation. Thefirst was the expanded role of RAC. On September24, 1976, its charter had been amended once more toprovide for additional expertise in the areas of botan y, plan t pathology, and tissue culture. More-over, its mem bership was in creased from 16 to 20 sothat four members would be “from other disciplinesor representatives of the general public. ” This wasthe first official provision for public representation

‘sIbid., pp. 52.Iafnterim Repof l of t he Federal Interagency Committee On flec0l7?-

binant DNA Research: Suggested Elements for Legislation, Mar. 15,1977, pp . 3.4.“[bid., pp. 9-10.‘“Ibid., pp. 11-15.q For a more complete discussion of the legislation, see footnote

19.

‘g.’’ Recombinant DNA Molecule Research, ” Congressional Re-

search Service, issue b rief No. IB 77024, upd ate o f Jan , 2, 1979.

although tw o nonscientists were already members.The number of nonscientists remained the sameuntil December 1978.

20Also, RAC’s resp onsibilities

were defined in greater detail, including the respon-sibility for reviewing large-scale experiments. N ever-theless, RAC continued formally at least to be “a tech-nical committee, established to look at a specificproblem.”

The second d evelopm ent was a grow ing belief among scientists that the r isks of the research w ereless than originally feared. This was based on the fol-lowing: 1) a letter from Roy Curtiss at the Universityof Alabama to the Director of NIH, explaining risk assessment experiments using Escherichia coli, fromwhich he concluded that the use of E. coli K-12 host-vectors posed n o dan ger to hum ans; 2) the conclu-sions of a committee of experts in infectious diseasesassembled by NIH in June 1977 in Falmouth, Mass.,that the alleged hazards of the research w ere un-substantiated; and 3) a prepublication report on ex-perimen ts showing that genetic recombination oc-curs naturally between lower and higher life forms,and suggesting that the rDNA techniqu e was not asnovel as presumed.

The third development affecting the legislationwas a concerted lobbying effort by scientists againstwhat they considered to be some of the overlyrestrictive provisions of the bills, especially S.

1217.21 22 23

The efforts included wide circulation of reports (including some in draft form) as soon as

a va i l a b l e , whi c h suppor t e d t he c onc l us i on t ha t

the research was less hazardous than original ly

supposed.By the end of 1977, the legislation was in limbo.

This si tuat ion continued in early 1978, al though

some hearings were held. On June 1, 1978, Senators

Kennedy, Javits, Nelson, Stevenson, Williams, and

Schweiker addressed a l e t t e r to HEW Secre ta ry

Joseph Califano, which acknowledged the likelihood

that legislation would not pass and urged that defi-

c ienc ies in the regula tory sys tem be addressedthrough executive action based on existing authority,

if that were to b e the case.

During Phase IV , NIH and its p arent organization,HEW (now DHHS), have attempted to operate in theregulatory vacuum left by the lack of legislation. Inresponse to th e consensus th at developed in 1977 on

~Wi l l i am Gar t l and, Director of the Office of Recombinant DN AActivities, NIH, personal communication, June 19, 1980.ZIB. Cu]] i ton , “Recombinant DNA Bills Derailed: COngre55 Still

Tryin g to P ass Law ,” Scien ce, vol. 199, Jan. 20, 1978. pp. 274-277.“D . Dickson, “Friends of D N A Fight Back,” Nature, vol. 272,

April 1978, pp. 664-665.Z SR . ~win, ,, Recombin ant DNA as a political pa~n,~’ ~e~ ~ien.

fist, VO]. 79, Sept. 7, 1978, pp. 672-674.

76-565 0 - 81 - 22



318 q Impacts of Applied Genetics-Micro-Organisms, Plants, and Animals

th e question of risk, RAC prop osed revisions to theGuidelines, which placed m ost experiments a t alower containment level. They were published forpublic comment in September 1977. * As with theoriginal Guidelines, public hearings were held in thecourse of a 2-day meeting of the Advisory Committeeto the Director in December 1977, in wh ich a d iverse

group of individuals and organizations were permit-ted to comment. However, at this point, HEW took amuch m ore active role in a situation th at had beenhandled almost entirely by NIH.

24

When RAC’s charter was renewed on June 30,

1978, Secretary Califano reserved the power to ap-point it s members instead of delegat ing it to theDirector of NIH as in the p ast.** And the n ew p ro-posed G uidelines, published in the Federal Registeron July 28, 1978, were accompanied by an introduc-tory statement by Secretary Califano announcing a60 day public comment period to be followed by apublic hearing before a departmental panel chairedby HEW General Counsel Peter Libassi. * * * The

Secretary was particularly interested in commen tson: new mechanisms to provide for future discre-tionary revision of the Guidelines; and the composi-tion of the various ad visory bodies, especially theRAC and the local Institutional Biosafety Committees(IBCS).

25

The p ub lic hearing called for by Secretary Cali-fano and held on September 15, 1978, was a sig-

nificant event in the history of Federal actions on therDNA issue. Testimony was heard from represent-atives of industry, labor, the research comm un ity,and public interest groups; more than 170 letters of comment were received and subsequently reviewed.As a result, the revised final Guidelines of December

22, 1978, were significantly rewritten to increasepublic participation in the decisionmaking process:

26

q Twenty percent of the members of the IBCs hadto represent th e general public and could h aveno connection with the institution.

q Most of the records of the IBCs had to b e pu blic-ly available.

q Shortly thereafter, in October 1977, the Final EnvironmentalImpact Statement for the 1976 Guidelines was published.

Z4D. Fredrickson, “A History of the Recombinant DNA Guide-

lines in the Un ited States,” Hecornbinmt DN A Technical Bulletin,

vol. 2, Ju]y 1979. p p. 87, 90.qq The statement providing for delegation of authority th at ac-

companied the updated Charter was not signed by Califano. Seealso, footnote 24.

* q q The other members of the HEW panel were Dr.Fredrickson, Julius Richmond, who was the Assistant Secretaryfor Health, and H enry Aaron, who was the Assistant Secretary forPlanning and Evaluation.=43 F.R. 33042, Ju ly 28, 1978.~estatement of Secretav Califano accompanying the revised

Guid elines, 43 F .R . 60080, Dec. 22 , 1978.

q Major actions, such as decisions to except other-wise prohibited experiments on a case-by-casebasis or to change the G uidelines, could b e madeonly on the advice of RAC and after public andFederal agency comment.

The increased public responsiveness of the IBC’swas crucial, since the revised Guidelines placed ma-

jor responsibility for compliance on them. This hadbeen proposed in th e July version and h ad not beenchanged by the hearings. * Califano also announcedhe would appoint 14 new members to the RAC, in-cluding people knowledgeable in fields such as law,pu blic policy, ethics, the environ men t, and p ub lichealth. Z’* q All of these changes were envisioned to“provide the opportunity for those concerned toraise any ethical issues posed by recombinant DNAresearch” an d to change the role of the RAC to “serveas the principal advisory body to the Director of NIHand the Secretary of HEW on recombinant DNAPoli cy .’’

28* * *

In add ition to b roadening pu blic participation,

Califano attempted to deal with a major limitation of the Federal response—the Guidelines did not coverprivate research, He directed the Food and Drug Ad-ministration (FDA) to take steps to require that anyfirm seeking approval of a product requiring the useof rDNA techniq ues in its development or manu-facture, demonstrate compliance with the Guidelinesfor the work done on that produ ct; an FDA n otice of its intention to propose such regulations accom-panied the revised Guid elines in the Federal Register.In addition, he requ ested the Environmental Protec-tion Agen cy (EPA) to review its regu latory authorityin that area. He believed if both agencies couldregulate research on products within their jurisdic-

tion, “virtually all recombinant DNA research in thiscountry would be brought under the requirementsof the revised guidelines. ’’

29* In the meantime, the

q As part of the revision process, HEW held a meeting in October1978 for IB C chairpersons in order to exchange in formation andexperiences gained under the 1976 Guidelines.

Z7 1 b i d .

q q This was implemented by an amendment to the RA C Charteron Dec. 28, 1978, which increased the membership to 25 and

changed the composition to the following categories: 1) at leasteight specialists in molecular biology or D N A research; 2) at leastsix specialists in other scientific fields; and 3) at least six personsknow ledgeable in law, public policy, the environmen t, and p ublicor occupational h ealth. In addition, the Charter was amend ed togrant non voting representation to representatives of various Fed-eral agencies.

Z s I b i d .

q * q The Charter was never amend ed to change or delete thefinal sentence of the “Purpose” section, which states, “This Corn.

mittee is a technical committee, established to look at a specificproblem.”

‘“Ibid., p. 60081.



Appendix I//-A—History of the Recombinant DNA Debate q 319

revised Guidelines provided, for the first time, forvoluntary registration of projects with NIH, in whichthe registrant would agree to abide only by the con-tainment standards of the Guidelines.

31

Other major changes were embodied in the newGu idelin es. Because of the consensu s that the ex-periments posed lower risks than originally thought,some types of experiments were exempted, whilecontainment levels were lowered for almost allothers. In order to provide greater flexibility, theseGuidelines permitted exceptions on a case-by-casebasis, and included procedures for their change on apiecemeal basis without going through the whole in-ternal process at HEW. For major changes, the pro-cedure was: 1) publication of the proposed changesin the Federal Register at least 30 days prior to a RACmeeting; 2) RAC consideration of the proposedchanges; and 3) publication in the Federal Register of the final decision of the Director, NIH. The standardfor all actions of the Director under the Guidelineswas “no significant risk to health or to the environ-merit. ”

32Lastly, the new Guidelines delegated project

approval to the IBCs.The problems posed by voluntary compliance and

commercialization have continued to be addressedby NIH. In a second major revision to the Guidelineson January 29, 1980, a section (Part VI ) was added tospecify procedures for voluntary compliance. * q O n

(cominuedfmm p. 318)

q Subsequently, Califano sent similar letters to the Secretaries of Agriculture [February 1979) and Labor (July 1979) requ estingthem to consider h ow their agencies’ authorities could be u sed torequire private sector rDNA research to comply with theGuidelines. qo

aoMinutes of the Interagency Committee on Recombin ant DNA

Research, p. 3, July 17, 1979, preprinted in Recombinant DNA Re- search, vol. 5, p. 132, et. seq.

alsec. IV.F.3, 1978 Guidelines.

3zSec. IV-E-l-b.

q “Several responses to the FDA notice had questioned the agen.cy’s legal authority to regulate private rDNA research. Conse.

quently , Dr. Fredrickson and Dr. Donald Kenned y, then Commis-sioner of Food and Drugs, developed a draft supplement to theGuidelines, specifying procedures for voluntary compliance by in.dus t ry . It was published for comment on Aug. 3, 1979 (44 F.R.45868) and incorporated a s part of th e prop osed revised Guide .

lines of November 30, 1979. (44 F.R. 69210, 69247).

April 11, 1980, NIH published Physical ContainmentRecommendations for Large Scale Uses of OrganismsContaining Recombinant DNA Molecules in the formof Draft Part VII to the Guidelines.

33Besides setting

large scale containment levels, this document recom-mend s that the institu tion: appoint a biological safetyofficer with specified duties; and establish a workerhealth surv eillance program for work requiring ahigh (P3) containment level. Finally, a more ad hoc re-quirement has been used since October 1979 for ap-provals of industrial requests for cultures up to 750liters (1); the ap provals were cond itioned on N IHdesignated observers being p ermitted b y the com-panies to inspect their facilities.

34At least one inspec-

tion has taken place.On November 21, 1980, NIH adopted the third ma-

jor revision to the Guidelines.35

It contained thesesignificant changes: institutions sponsoring theresearch are no longer required to register theirprojects with NIH pursuant to an informational docu-ment called a Memorandum of Understanding (MUA)whenever the containment levels are specified in theGuidelines; and NIH will no longer review IBC deci-sions on experiments for which containment levelsare specified in the Guidelines.

On November 21, 1980, NIH also promulgatedrevised application procedures for large-scale pro-posals. The app lication must in clude the following in-formation: 1) the registration document submitted tothe local IBC; 2) the reason for wanting to exceed the10-1 limit; 3) evidence that the rDN A to be used wasrigorously characterized and free of harmful se-quences; and 4) specification of the large-scale con-tainment level proposed to be used as defined in theNIH Physical Containment Recommendations of April 11, 1980.

In addition to addin g part VI to the Guidelines, the most signifi-cant change in the January 1980 Guidelines was the ad dition of sec. 111-0, which permitted most experiments using E. coli K-12host-vector systems to be done at the lowest containment levels.

s345 F.R. 24%8, Apr. 11, 1980.

3444 F.R. 69251, NOV. 30, 1979.3345 F.R. 77372, Nov. 21, 1980.



Appendix III-B

Co n s t it u t io n a l C on s t r a in t s

o n R e g u l a t i o n

Under the checks and balances of our system of government, the Constitution, as ultimately inter-preted by th e Sup reme Court, requires certain pro-cedural and substantive standards to be met by stat-utory or other regulation imposed u pon an activity.These requirements depend on the nature of the ac-tivity involved. In the present case, it will be useful toconsider first the regu lation of basic research andthen the regulation of technological applications,such as the p rodu ction of pharm aceuticals by usinggenetic engineering methods.

Research

With respect to research, the fundamental ques-tion is what limitations, if any, may be placed on thesearch for scientific knowledge. The primary appli-cable constitutional provision is the first amendment,which has been broadly interpreted by the SupremeCourt to severely limit intrusion by the Governmenton all forms of expression.

123Another constitutional

safeguard, known as equal protection, is secondarilyinvolved.

If the Supreme Court were to recognize a right of scientific inquiry, its boundaries would not exceedthose for freedom of expression.

4There is disagree-

ment among commentators on this issue concerningthe boun daries of the first amendmen ts and certain-ly disagreement on the application of generally ac-cepted principles to particular cases. Moreover,there have been n o judicial decisions dealing with th eprecise issue at hand. However, it is possible to out-line general prin ciples derived from jud icial deci-sions interpreting the first amendment, and indicatehow th ey might be applied b y the courts to attemptsto regulate genetic research.

There are very few limitations on th e written orspoken word. The prohib itions against obscenity or“fighting words”* clearly would b e inapp licable here.

‘Harold P. Green, “The Soundaries of Scientific Freedom” Regulation of

Scientt~ic Inquiry: ticietal Concerns With Research, Keith M. Wulff (cd.)

(Washington, D. C.: AAAS, 1979), pp. 139-143.

aThomas 1. Emerson, “The Constitution and Regulation of Research,” fteg-

u/ation o~scientt~ic inquiry: Societal Concerns With Reseamh, Keith M. Wulff

(cd.) (Washington, D. C.: AAAS, 1979), pp. 129-137.

‘John A. Robertson, “The Scientists’ Right to Research: A Constitutional

Analysis,” Soufhern Ca/~j’ornia Law Review 5 1:1203, September 1978.4Green, op. cit., p. 140.

‘Emerson, op. cit., pp. 131-134.

“’Fighting words” are those provoking violent reaction or imminent

disorder.

For many years, the Supreme Court has conceptual-ized the right of free expression in terms of a market-place of ideas—through the open and full discussionof all ideas and related information, the valuable,valid, or useful ones will be accepted by society,while the ridiculous or even dangerous ones will beso demonstrated and discarded. This is a consensualprocess; no person, group, or institution has suffi-cient wisdom to prejud ge ideas and d eny themadm ittance to that intellectual m arketplace, even if they threaten fund amental cultural values, for suchvalues, if worthwh ile, will survive. Under th is con-cept, scientists wou ld certainly have virtually un re-strained freedom to think, speak, and write.

Difficulties arise with actions, such as experimen-tation, which may be essential to the implementationof freedom of expression. Recent Supreme Courtcases have recognized a limited protected interest of the med ia to gather information as an essential ad-

junct to freedom of publication. By analogy, it maybe argued that scientists would also be protected intheir research, as a necessary adjunct to freedom of expression. On the other hand , the informationgathering cases usually involve access to Govern-ment facilities, such as courtrooms or prisons. Theyare based on th e principle that actions by the Gov-ernment should be open to pu blic scrutiny—a con-cept not directly applicable to the present issue.

More importantly, the Court has long recognizedthat actions related to expression can be regulatedand that regulation may increase with the degree of the action’s impact on people or the environment.The Court would probably apply what has beencalled a structured balancing test;

6i.e., regulation

would be deemed valid only when the Governmentsustains the bu rden of p roving: 1) that there are“compelling reasons” for the regulation; and 2) thatthe objective cannot be achieved by “less drasticmeans, ” i.e., by more narrowly drafted regulationshaving less impact on first amendment rights.

The second part of the test is fairly straightfor-ward. Governmental restrictions must be kept to a

minimum. E.g., where possible, they should be reg-ulatory rather than prohibitory, temporary ratherthan permanent, involve the least burden, and so on.

The d ifficult par t of this test lies in determining

‘Ibid., p. 134.

320



Apppendix III-B—Constitutional Constraints on Regulation q 321

what is a compelling reason. The protection of healthor the en vironment is the m ost clearly acceptablereason for regulation. In addition, the protection of individual rights and personal dignity is generallyconsidered an acceptable reason. E.g., the NationalResearch Act

7requires that all biomedical and be-

havioral research involving human subjects sup-

ported u nd er the Pub lic Health Service Act be re-viewed by an Institutional Review Board in order toprotect the rights and welfare of the subjects.

The above d iscussion relates to protection fromphysical risks due to the process of research. Couldthe Government regulate or forbid experimentationsolely because the p roduct (knowledge) threatenscultural values or other intangibles such as the genet-ic inheritance of mankind? Religious or philosophicalobjections to research, based solely on the rationalethat there are some things mankind should notknow, conflict with the basic principles of freedomof expression and would not be sufficient reason onconstitutional grounds to justify regulation. Even if

the rationale underlying this objection were expand-ed to include situations where knowledge threatensfundamen tal cultural values about the n ature of man, control of research for such a reason probablywould not be constitutionally permissible. The ra-tionale would again conflict with the m arketplace of ideas concept that is central to freedom of expres-sion. However, what if the knowledge were to pro-vide the means to alter the human species in such away that the physical, psychological, and emotionalessence of what it is to be human could be changed?No p recedent exists to provide guidance in d etermin-ing an answer. Were the situation to arise, theSupreme Court might fashion another limitation on

the concept of free expression in the same w ay itdeveloped th e obscenity or “fighting words” d oc-trines.

The discussion thus far has had as its premise adirect regulatory app roach to research. There is amore indirect approach, wh ich wou ld be constitu-tionally permissible and could accomplish much of what direct regulation might attempt, including pre-vention of the acquisition of some forms of knowl-edge. This is the use of the funding power. Thelifeblood of modern science in the United States isthe Federal grant system. Yet it is generally agreedthat Government has no constitutional du ty to fundscientific researche

sThis is a benefit voluntarily pro-

vided to which many k inds of conditions may be at-tached. The only constitutional limitation on such anapproach would be the concept of equal protection—any restrictions must apply to all or must not be ap-

TPublic Law 93-348 (1974), 42 U.S. C. S289 1-3

‘Green, op. cit., p. 141.

plied in a discriminatory way without compellingreasons.

Congress could therefore, mandate by law thatcertain kin ds of research n ot be fun ded or be con-ducted in certain ways. An example is the NationalResearch Act, discussed previously. However, thisapp roach may h ave some serious p ractical limita-

tions because of the difficulty of determining whichmolecular biological research might lead to the pro-scribed knowledge. Much discretion would have tobe left to the fund ing agency, which is likely to be un -sympathetic or even hostile to such an approach, if itviews its primary mission as fostering research.

A p p l i ca t i o n s a n d p r o d u c t s

Although fears have been expressed that currentgenetic techn ologies may lead to app lications thatwould be detrimen tal, no one can reasonably con-clude, at the present time, that this will actually oc-cur. For this reason, the most constitutionally p er-missible approach in all probability will be to regu-

late the app lications of the science. In su ch situa-tions, whatever harms occur tend to be more tangi-ble and the governmental interests, therefore, moreclearly defined. Moreover, since fundamental con-stitutional rights are generally not involved, statutesand regulations are subjected to a lower level of scrutiny by the Federal courts.

The constitutional authority for Federal regulationof the applications of technologies such as genetic en-gineering lies in the commerce clause, article I, sec-tion 8 of the Constitution, which grants Congress thepower “To Regulate Commerce with foreign Nations,and among th e Several States. ” In contrast to sit-uations involving fun damental rights, the Supreme

Court has interpreted this clause as giving Congressextremely broad authority to regulate an y activity inany way connected with commerce. It has been vir-tually imp ossible for Congress not to find some con-nection acceptable to the courts between commerceand the goals of a particular piece of legislation. * Thestandard of review of such legislation by the Federalcourts is to determine if it bears a rational re-lationship to a valid legislative pu rpose. If so, theCourt will uphold the legislation and will not secondguess the legislators. This stand ard of review rec-ognizes that a statute results from the balancing of comp eting interests and policies by the branch of Government created to function in that manner.

*See Wickard v. Filburn, 317 U.S. Ill 0 9 4 2 ) in which the Supreme Court up-

held civil penalties for violation of acreage allotments established by the

Agricultural Adjustment Act of 1938, covering the amount of wheat that in-

dividual farmers could plant, even if the wheat was intended for selfcon-

sumption. The rationale was that even though the individual farmer’s wheat

had no measurable impact on interstate commerce, Congress could prop-

erly determine that all wheat of this category, if exempted from regulation,

could undercut the purpose of the Act, which was to increase the price

farmers received for their various crops.



Appendix III-C

In for m a t i on on In t e r n a t i on a l

G u i d e l i n e s f o r R e c o m b i n a n t D N A

The following information is based largely on in-ternational surveys undertaken by The Committeeon Genetic Experimentation of the InternationalCouncil of Scientific Unions reported as of July1979.’

I . Nations that had established guidelines for conduct of rDNA research or were using the guidelines of other nations:

AustraliaBelgiumBrazilBulgariaC an ad a

CzechoslovakiaDenmark German Democratic

RepublicFederal Republic of

GermanyFinlandFranceHungaryIsrael

ItalyJapanMexicoNetherlandsNew Zealand

NorwayPolandSouth AfricaSwedenSwitzerlandTaiwanUnited KingdomUnited StatesU.S.S.R.Yugoslavia

II . Nations that had not established guidelines or had not responded with updated information:

Country Yes N o

AustriaGhanaIndiaIranJamaicaKoreaNigeriaSingaporeSri LankaSudanTurkey

x

xx

x

xx

x

xx

x

x

111. Nations that had drafted their own guidelines:

Canada JapanFederal Republic of United Kingdom

‘Report to COGENE ftmm the working gruup on Recombinant DNA Guide-

lines, May 1980.

Germany United StatesFrance U.S.S.R.Italy

IV. Nations that had modified the guidelines of other,indicated, countries:

Au stralia (UK, U. S.)Belgium (UK, U. S.)Brazil (U. S.)Bulgaria (U. S. S. R., U. S.)Czechoslovakia (U. S. S. R.,

U. S., Fed. Rep . Ge r.)Denmark (UK)East German Democratic

Republic (UK, U. S.,Nether lands )

Finland (U.S. mainly)Hu ngary (U. S.)

Mexico (U. S.)Netherlan ds (U. S.)New ZealandNorw ay (U. S.)Poland (U. S.)South Africa (U. S.)Sweden (U.S)Switzerland (U. S.)

Taiw an (U. S., UK)Yugoslavia

(European ScienceFoundation)

V. Nations in which entirely voluntary guidelines have

been adopted:

Finland

VI. Natio ns w ith guidelines that are enforceablethrough control of research funding:

Australia” JapanC an ad a NetherlandsCzech oslovak ia Norway

Denmark South AfricaFederal Republic of S wed en

Germany’ SwitzerlandFrance Taiwan

e

German Democratic United Kingdomf

Republic United States

a’isubmissions may be made directly to the Academy of Science or

through a granting agency. In the latter case, it is a requirement for the ap-

plicant to observe the recommendations of the Academy’s Standing Com-

mittee if the agency makes a grant for the work. Otherwise, the guidelines

are voluntary with the worker required to make an annual report on prog-

ress, or more frequently if conditions of the experiment (such as volumes)

are changed appreciably.”b’~ntrol through Academy of Sciences and Ministry of Health.”

c~veral research organ~tions requi re MC e iV e r s of grants to apply ‘he

NIH guidelines until their own national guidelines are completed.dThe Netherlands Organization for the Advancement of Pure Research

will only subsidize projects which have been given the committee’s conaent.e!(waiting fo

rre5pon5e from National Advisory CCJmmittee. ”

f“Notification of proposals to GMAG became compulsory August 1, 1978.

In addition, funding bodies require, as a condition of funding, GMAG’s ad-vice to be sought and followed. ”

322



Appendix I//-C—Information on International Guidelines for Recombinant DNA q 323

VII. Nat ions in w hich guidelines are legally enforceable:

Hungary

U.S.S.R.

Finland. . . . . . . “At p rese nt , the guidel ines a r e

South Africa. .

entirely voluntary, but in the near

future, the intent ion is to includethem in the law of infectious dis-

eases when they will become legally

enforceable. ”

“At present the guidelines are not

legally enforceable. They will only

become so if regulations under the

existing Health Act of 1977 and the

Animal D iseases and Parasites Act of

1956 ar e promulgated; and none arein tended at p resent . ”

United Kingdom “The regulation to notify GMAGdoes not strictly mean that theWilliams Guidelines themselves are

legally enforceable. But, under theHealth and Safety at Work Act(within which the Regulations wereintrod uced), it is expected th at ac-count will be taken of the relevantCodes of Practice and the advicegiven by GMAG .”

VIII. Nat ions in w hich observance of the guidelines ismonit ored by a nationally -directed mechanism:

Australia Norway

Czechoslovakia South Africa

German Democratic S wed enRepublic United Kingdom

France United StatesHungary U.S.S.R.Japan Yugoslavia

IX. Nations in which a license or other authorization for recombinant DNA activity is granted:

—to an institution: U.S.S.R.—to an indivdual laboratory: Hungary, Czechoslo-

vakia—to an individual scientist: Australia, Canada, Ger-

man Democratic Republic, Federal Republic of Germany, Finland, France, Japan, Norway, SouthAfrica, Sweden, United Kingdom”, United Statesand U.S.S.R.

Netherlands: “There are gentlemen’s agreements,signed by the ind ividual scientist, the institutionand the Committee.” The reports of the Committeealso recommend legislation that will require regis-tration of research projects in this field and makebind ing the guidelines and sup ervision of theirobservance. (Report of t he Committee in Charge of

the Control on Genetic Manipulation, Amsterdam,March 1977, p. 54.)

Bulgaria, Switzerland: None of the above.Taiwan: No response.

them in the context of information about the ‘centre’ in which the work is to

go on.”

X. Nations in which special provisions for agriculture and/or industrial research and applications have been made:

Czechoslovakia. “10 liter maximum volum e of thecu ltu re con tain in g recombinantDN A”

German

Democratic

Republic. . . . . “The GDR Gu idelines will be com-pulsory for industrial and agricul-

tural applications. 10-liter maximumdeviations may b e allowed b y theMinister of Health if suggested by

the Committee. ”Federal Repub licof Germany . . “Specification of containment of

plants”France . . . . . . . “Industry, maximum volume of cell

culture is set at 10 liters”Norway . . . . . . “The G uid elines cover b oth agri-

culture and industry. Application of recombinant DNA research outsidean app roved laboratory is prohib-ited. Otherwise the Committee fol-lows the NIH Guidelines .”

United Kingdom “Agriculture, industry; see WilliamsReport, paragraphs 1.3, 2.7, 5.13and appendix II, section 34. ”

United States . . “Agriculture. NIH Guidelines pro-

U.S.S.R. . . . . . .

Other

respondents, .

vide containment levels for cloningtotal plant D NA, plant virus DNAand plant organelle DNA in E. coliK-12, and provide general guidancefor the use of plant host-vector sys-tems. 10 liter maximum. A proposedSupplemen t to the Guidelines forvoluntary compliance by the privatesector is under consideration byRAC. Development of a monographfor large-scale applications has beenproposed.”“Guidelines are compulsory for in-dustrial and agricultural applica-tions. 10 liter maximum. Deviationis allowed by the Recombinant DNACommission. ”

N o



324 q impacts of Applied Genetics—Micro-Organisms, Plants, and Animals

XI. Number of laboratories currently engaged in recombinant DNA activities:

Country Any labs? How many?

Australia. . . . . . . . . . . .Austria . . . . . . . . . . . . .Belgium. . . . . . . . . . . . .

Brazil. . . . . . . . . . . . . . .Bulgaria. . . . . . . . . ... ,Canada . . . . . . . . . . . . .Czech oslov ak ia ., . . . . .Denmark. . . . . . . . . . . .German Democratic

Republic . . . . . . . . . . .Federal Repub lic

of G erm any. . . . . . . . .Finland ., . . . . . . . . . . .France. ., . . . . . . . . . . .Ghana . . . . . . . . . . . . . .Hungary . . . . . . . . . . . .India . . . . . . . . . . . . . . .

Iran. . . . . . . . . . . . . . . .Israel. . . . . . . . . . . . . . .Jamaica. . . . . . . . . . . . .Japan . . . . . . . . . . . . . .Korea . . . . . . . . . . . . . .Netherlands . . . . . . . . .New Zealan d. . . . . . . . .Nigeria . . . . . . . . . . . . .Norway . . . . . . . . . . . . .Phi lip pines . . . . . . . . . .Poland . . . . . . . . . . . . . .Singapore . . . . . . . . . . .Sou th Africa . . . . . . . . .Sri Lanka. . . . . . . . . . . .

Sudan . . . . . . . . . . . . . .Sweden. . . . . . . . . . . . .Switzerland. ., . . . . . . .Taiwan . . . . . . . . . . . . .Turkey . . . . . . . . . . . . .United Kingd om . . . . . .United States.. . . . . . . .U.S.S.R . . . . . . . . . . . . . .Yugos lav ia. . . . . . . . . . .

ye sno

a

ye sa

ye sye s

a

yesyesyes a

ye s

yesye sye s

a

n oa

yesa

n oa

n o

a

yesa

n oa

yesn o

a

ye syesn o

ye sn o

a

yesa

n oa

yesa

n oa

n o

a

yesa

yesa

ye sn o

a

ye syes

a

yes a

yesb

16

6

5no response

10-153

several

5

10-203(3-4 planned)

12

1-2

1

35

7

2

not stated

3

3

2

18

2

45

50

6

4

XII.Countries in which specific training for workers and safety officers in recombinant DNA activities is required by the guidelines:

Country Yes N o Other

Austral ia . . . . .Bulgar ia. . . . . .

Can ada . . . . . . .Czechoslovakia.German

DemocraticRepublic . . . . .

Federal Republicof Germany . .

Finland. . , . . . .France . . . . . . .Hu ngary. . . . . .Japan. ..., . . .Netherlands. . .Norway . . . . . .South Africa. . .

Sweden . . . . . .Switzerland . . .Taiw an . . . . . . .United KingdomUnited States . .U.S.S.R. . . . . . .Yugos lavia . . . .

a

x

xX

b

xc

Xd

x

Xe

x

X g

x

x

x

x

Xf

x

“recommended”“recommended”

Xh

no response

Other respondents: no or no response to question.

aAustra]ia: URequireexpefii~ through Biosafety COmmiWW.”

bczechoslovakia: ~spific training is recommended.”

c~rman Democratic Republic: “Training courses are organized by the

Committees in cooperation with Akademie fur Arztliche Fortbildungder

DDR.”dFederal Republic oftkrmany: “Experience as required by law on the

control ofcommunicable diseases.”eNether]ands: “Thescientistss hould be trained in microbiology.”

fNorway: The Committee certifies training and expertise of personnel are

adequate. ”

gunited I ( i n gdo r n : “De t a i ] S of training are required; the employer is legally

obliged to provide suitable training. ”hunited States: ‘“SFific training not required. However, local biohazards

committees are required to certify to the NIH that the training and expertise

of the personnel are adequate. ”

a~wd on replies f rom previous Questionnaires.

bln preparation.



Appendix III-C—lnformation on international Guidelines for Recombinant DNA q 325

X111. Countries in which the guidelines are applicable only to biological agents containing recombinant DNA, or also cover the recombinant DNA molecules themselves:

Only tobiological Also recombinant

Country agents DNA moleculesAustralia. . . . . . . .

Bulgaria . . . . . . . .

Canada. . . . . . . . . (a)

Czechoslovakia . .

GermanDemocraticRep ubl ic. . . . . . . x

Federal Republic of Germ any. . . . . . . x

Finlan d . . . . . . . . .France . . . . . . . . . x

Japan . . . . . . . . . . xNeth erlands . . . . .

New Zealand. . . . .Norway . . . . . . . .Sou th Africa. . . . .Sweden . . . . . . . . .Switzerl and . . . . .Taiwan. . . . . . . . . xUnited Kingdom . .United States . . . .U. S. S. R.. . . . . . . . .

x

(a)x

x

x

xx

x

x

x

x

X b

x

a(;uidelines appty to a]i, but containment is not required for naked DNA.

b’The [guidelines-apply to recombinant DNA experiments that are not ex-

empt under Section I-E of the Guidelines. Recombinant DNA molecules that

are not in organisms or viruses are exempt from the Guidelines (I-E-l ).”

XIV. Groups/Comm itt ees responsible for carryin g out monitoring of containment procedures:

Country Group

Australia. . . . . . . . Institu tional Biosafety Comm it-

tees.

Bulgaria . . . . . . . . National Committee

Canada . . . . . . . . . “Un iversity a n d M e d i c al R e-

search Council Biohazards Com-

mittees”Czechoslovakia. . . “Under consideration of the Na-

tional Institutes of Public Health.”German

DemocraticRep ub lic . . . . . . . “Monitoring is carried out by

local Biosafety Officers, who arerepresentatives of the Committeein their institution s. ”

Federal Republic of Germany . . . . . . . Officers for Biological Safety

monitor the health of em ployees

and comp liance at laboratories;ZKBS (Zentrale Commission fur

die biologische Sicherheit) hasoverall responsibility.

France . . . . . . . . . “Local safety committees”Hu ngary. ... , . . . “National Institu tes of Pub lic

Heal th”

Japan ... , . . . . . . “Principal Investigator and SafetyOfficer”

Neth erland s . . . . . “Site Inspection Commission”New Zealand. . . . . “Local controlling Committees

are charged with monitoringo b s e r v a n c e o f Guidelines.Biological Safety Officers are ap-po in ted to t ake im m edia t eresponsibility.”

Norway . . . . . . . . “Physical containm ent: Norwe-gian National Institute of PublicHealth. Biological containment:Committee. ”

South Africa. . . . . “Above P3, Biosafety Committee

of Ins t i t u t e involved andSAGENE. Below P3, SAGENEonly.”

Swed en . . . . . . . . . Not applicable.Switzerland . . . . . “At the responsibility of either

the ind ividual investigator or alocal biohazards committee.”

Taiwan. . . . . . . . . No responseUnited Kingdom . . The Health an d Safety ExecutiveUnited States . . . . “Observance of containment is to

be monitored by biohazards com-mittees located in institutions inwhich the research is conducted.Effectiveness of containment

procedures is to be monitored bythe principal investigator who isto report problems to the N IH.”

U. S. S. R.. . . . . . . . . “Local biosafety commission,State Sanitary Inspection controlgroup of Recombinan t DNA Com-mission.




xv . Countries in which the guidelines apply to all gene

combinations instructed by cell-free methods, or only to molecules containing combinations of genes from different species:

MoleculesAll gene com- containing

binations con- combinations of s tructed by ce ll - genes f romCountry free methods different species

Austral ia. . . . . . . .Canada. . . . . . . . .Czechoslovakia. . .German

DemocraticRep ubl ic. . . . . . .

Federal Republic of Germ any. . . . . . .

Finlan d. . . . . . . . .France . . . , . . . . .Japan . . . . . . . . . .

Neth erlands . . . . .New Zealand. . . . .Norway . . . . . . . .Sou th A frica . . . . .Sweden. . . . . . . . .Switzerl and . . . . .Taiwan. . . . , . . . .United Kingdom . .United States . . . .U. S. S. R.. . . . . . . . .

x

x

x

x

X a

xx

x

Xb

x

x

x

x

x

x

aFederal Republic of Germany Selfwloning experiments involving non-

pathogenic donors and hosts shall be reported to ZKBS.bNetherlands “The definition of recombinant DNA has recedy been

modified and includes the insertion of chemically synthesized DNA mole-

cules into a vector. ”cunited Kingdom “The G~u@s provision] interpretation Of their Own re-

mit is that they are concerned with work involving genetic manipulation,

defined for these purposes as: the formation of new combinations of her-

itable materials by the insertion of nucleic acid molecules, produced by

whatever means outside the cell, into any virus, bacterial plasmid, or other

vector sbystem so as to allow their incorporation into a host organism in

which they do not naturally occur but in which they are capable of con-

tinued propagation.”

XVI. Countries in which the guidelines restrict the intentional dissemination into the environment of biological agents containing recombinant DNA:

All respondents . .Austral ia. . . . . . . .German

DemocraticRep ub lic . . . . . . .

Yes a

Not explicity so

“Exceptions have to be discussedby the Committee and requirespecial permission by th e Minis-ter of Health.”

New Zealand. . . . . “Yes, with the approval of theNational Committee.”

United Kingdom . . “The question h as not arisen.”Other respondents No

aAr@ there anv circumstances under which such dissemination ca n b e c a r -

ried out?

XVII. Countries in which the guidelines are restricted to recombinant DNA activities or also cover other areas of genetic experimentation:

Recombinant Other areas of D N A genetic

Country activities experimentation

Australia. ... , ., .Bulgaria . . . . . . . .Can ad a . . . . . . . . .Czechoslovakia. . .German

DemocraticRep ub lic . . . . . . .

Federal Republic of Germ any. . . . . . .

Finland. . . . . . . . .France . . . . . . . . .Hungary. . . . . . . .Japan . . . . . . . . . .Neth erlands . . . . .New Zealand. . . . .Norway . . . . . . . .Sou th A frica. . . . .Swed en . . . . . . . . .Switzerland ... , .United Kingdom . .United States . . . .U. S. S. R.. . . . . . . . .

xa

x

Xb

x

x

x

x

x

x

x

x

x

xx

x

x

x

X C

Xd

a“At present, the terms of reference of the Academy Committee refer

only to in vitru experiments (i.e., the use of restriction enzymes and ligases).

An ad hoc Academy Committee is about to investigate in vivo experimenta-

tion, with the following terms of reference:

1. Examine whether, other than by using the technique of in v i t r u re-

combinant DNA construction, new hybrid nucleic acid molecules can be

produced that are potentially dangerous to humans, animals, or plants.

In so doing, the committee should give particular attention to the follow-

ing possibilities:

—The use of mixed infections involving human or animal viruses, or the

use of bacteria or fungi.

–The introduction of foreign DNA into plants and the production of

new plant pathogens.

2. Consider whether there are certain classes of viral pathogens (e.g.,

polio) on which experimentation should not be carried out unless a spe-

cial need is demonstrated.” ‘

b“work with animal viruses and cells”C“i.e., cell fusion with approval of National Committee”

d“other c]osely related areas are also covered.”



Appendix //l-C—Information on International Guidelines for Recombinant DNA . 327

XVIII. Countries in which the recombinant DNA ad-visory committee includes public representatives as well as scientists:

Country Yes N o

Australia. . . . . . . .

Bulgaria . . . . . . . .

Canada . . . . . . . . .Czechoslovakia. . .

Denmark . . . . . . .

German

Democratic

Republic . . . . . . .

Federal Repub lic of

Germany. . ,

Finland. . . . .

France . . . .,

Hungary. . . .

Italy . . . . . . .

Japan . . . . . .

Netherlands .

New Zealand.Norway . . . .

South Africa .Sweden. . . . .

Switzerland .

Taiwan. . . . .

. . . .

. . . .

. . . .

. . . .

... ,

. . . .

.,. .

. . . .

. . . .

. . . .

. . . .

. . . .

. . . .

United Kingdom . .

United States . . . .

U. S. S. R.. . . . . . . . .

x

x

x

x

x

Finland. . . . . . . . .

x

x

x

x

x

x

x

x

x

x

x

x

x

x

x

x

x

Composition of DNA advisory committees is as fol-lows:Australia. . . . . . . .

Canada . . . . . . . . .

Czechoslovakia. . .

Denmark . . . . . . .

GermanDemocraticRep ub lic . . . . . . .

8 scientists5 laymen (1 lawyer, 1 bu siness-

man, 3 generalists); 6 scientists (2M.D.s, 3 virologists/cancer sp e-cialists, 1 recomb inant DN A spe-cialist)6 memb ers representing molec-ular biology, genetics, microbiol-ogy, medicine9 scientists and administrativerepresentatives.

3 geneticists, 1 biochemist, 2 bac-teriologists, 2 virologist, 1 jurist,1 representative of trade union

of GDR.Federal Republic of Germany. . . . . . . 4 experts working in the field of

recombinant DNA research; 4 ex-perts who, though n ot workingin the field of recombinant D NA

France . . . . . . . . .

Hungary. , . . . . . .

Italy . . . . . . . . . . .

Japan . . . . . . . . . .

Netherlands . . . . .

New Zealand. . . . .

Norway . . . . . . . .

Sou th A frica . . . . .

Sweden. . . . . . . . .

research, p ossess specific knowl-edge in the implementation of safety measures in biological re-search work, particularly how-ever in microbiology, cytobiolo-gy, or hygiene and, in addition, 4outstanding individuals, for ex-

ample from the trade un ions, in-dustry, and the research-promot-ing organizations.27 members: 6 molecular biol-ogy, 3 genetics, 3 microbiology, 1virology, 1 plant physiology, 3 in-fectious diseases, 3 epidemiology,2 enteric bacteria, 1 cell cultures,3 public health, 1 occupationalhealth .13 mem be rs, 4 obs ervers, 1 sec-retaryScientists8 molecular biologists, 4 micro-

biologists, 1 civil servant (HealthMinistry).(Combines both Steering Com-mittee and Advisory Group): 7 re-combinant DNA scientists, 7 sci-entists in other fields, 6 special-ists in medicine and biohazards, 2lawyers, 2 specialists in physicalcontainment, 3 public represent-atives.14 scientists representing genet-ics, molecular biology, bacteriolo-gy, virology, botany, m edicine,ethics and social aspects of health

and health-care. To be add ed: acommittee composed of scientistsand representatives of ind ustryand trade unions.

1 molecular biologist, 1 microbialgeneticist, 1 virologist, 1 botanist

(molecular biologist), 1 hu mangeneticist (medically qualified).3 biochemists, 2 medicine, 1 vet-erinary medicine, 1 lawyer, 1artist.

On e each from: Council for Sci-entific and In du strial Research,

Med ical Research Coun cil, De-partment of Health, Departmentof Agricultural Technical Serv-ices. Three from universities,pub lic and legal professions.

No response




Switzerland ... , . 12 members representing medi- ogy: 2, Plant Genetics: 2, Law: 2,cin e, m icrob iology, m olecu lar Environmental Concerns, Lab-biology, antibiotics, industry, oratory Technician, Infectiousuniversity management, and 7 Diseases, Occupational Health,governm enta l depar tm en ta l Education: 1 each.assessors. U. S. S. R.. .. ......8 scientists

United States . . . . Molecular biology: 6, Molecular Yugoslavia .. ....3 geneticists

Genetics: 5, Ethics: 3, Microbiol-



Appendix IV

P l a n n i n g W o r k s h o p P a r t i c i p a n t s ,

O t h e r C o n t r a c t o r s a n d C o n t r i b u t o r s ,

a n d A c k n o w l e d g m e n t s

P l a n n i n g w o r k s h o p p a r t i c i p a n t s

Philip Bereano

University of Washington

Ralph Hardy

E. L Du Pent de Nemours & Co., Inc.Patricia King

Georgetown University Law Center

Charles LewisU.S. Department of Agriculture

Herman Lewis

National Science Foundation

Pamela Lippe

Friends of the Earth

Victor McKusick

Johns Hopkins University Medical School

Elena O. Nightingale

Institute of Medicine

Gilbert Omenn

Office of Science and Technology Policy

Donna Parratt

Congressional Research Service

Walter Shropshire

Smithsonian Radiation Biology Laboratory

Leroy Walters

The Kennedy Institute

Oth er co n t r a c to r s a n d co n t r ib u to r s

Betsy Amin -ArsalaRichard J. Auchus

Massachusetts Institute of TechnologyFred Bergmann

National Institutes of HealthCharles L. Cooney

Massachusetts Institute of TechnologyRichard CurtinRobert A. Cuzick

Massachusetts Institute of TechnologyRoslyn Dauber

Arnold L. DemainMassachu setts Institute of Technology

Richard B. EmmittF. Eberstadt & Co.

Emanuel EpsteinUniversity of California, Davis

Robert F. FleischakerMassachusetts Institute of Technology

Odelia FunkeGeorge E. Garrison

F. Eberstadt & Co.Reinaldo F. Gomez

Massachusetts Institute of TechnologyJohn HamiltonNeal F. JensenScott A. King

F. Eberstadt & Co.Harvey Lodish

Massachusetts Institute of TechnologyL. D. Nyhart

Massachusetts Institute of TechnologyChoKyun Rha

Massachusetts Institute of TechnologyWilliam ScanlonAndrew Schmitz

University of California, BerkeleyMichael J. Shodell

The Sterling-Hobe Corp.David Tse

Massachu setts Institute of TechnologyJames Welsh

Montana State UniversityGeorge Whiteside

Massachusetts Institute of Technology

Bernard Wolnak an d AssociatesNancy Woods

OTA intern

A c k n o w l e d g m e n t s

A large number of in dividuals provided valuableadvice and assistance to OTA du ring this assessment.In particular, we would like to thank the followingpeople:John Adams

Pharmaceutical Manufacturers AssociationRupert Amann

Colorado State University

William Amen, Jr.Cetus Corp.

Daniel Azarnoff Searle Laboratories

A. L. BarrWest Virginia University

K. J. BetteridgeAnimal Diseases Research Institute

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Jerome BirnbaumMerck, Sharp & Dohme Research Laboratories

Gerald BjorgeU.S. Patent and Trademark Office

Hugh BollingerPlant Resources Institute

G. Eric Bradford

University of CaliforniaRobert Brackett

Parke, Davis & Co.Robert Byrnes

Genentech, Inc.Daniel Callahan

The Hastings CenterAlexander M. Capron

President’s Commission for the Study of EthicalProblems in Medicine and Biomedical andBehavioral Research

Robert ChurchUniversity of Calgary

H. Wallis Clark

University of CaliforniaGail Cooper

Environmental Protection AgencyJoseph P. Dailey

Revlon Health Care GroupFrank Dickinson

Agricultural Research CenterDonald R. Dunner

Finnegan, Henderson, Farabow, Garrett &Dunner

Roger B. DworkinIndiana University School of Law

Richard P. ElanderBristol-Myers Co.

Peter ElsdenColorado State University

Haim ErderUniversity of Pennsylvania

James F. EvansPennsylvania Embryo Transfer Service

Kenneth EvansPlant Variety Protection Office

Richard FaustHoffman-La Roche, Inc.

Herman FinkeSterling Systems

Robert H. FooteCornell University

Orrie M. FriedmanCollaborative Research, Inc.

William J. Gartland , Jr.National Institutes of H ealth

Kenneth GoertzenSeed Research, Inc.

Michael GoldbergFood and Drug Administration

Maxwell GordonBristol Laboratories

Lorance L. GreenleeKeil and Witherspoon

Ralph Hardy

E. I. Du Pent de Nemours and Co,, Inc.W. C. D. Hare

Animal Diseases Research InstitutePaul Harvey

U.S. Department of AgricultureHarold W. Hawk

Agricultural Research CenterRichard L. Hinman

Pfizer, Inc.Peter Barton Hutt

Covington & BurlingE. Keith Inskeep

West Virginia UniversityIrving Johnson

tiny Research LaboratoriesCharles Kiddy

Agricultural Research CenterThomas D. Kiley

Genentech, Inc.Carole Kitti

National Science FoundationDuane C. Kraemer

Texas A&M UniversitySheldon Krimsky

Tufts UniversityW. W. Lampeter

Lehn und VersuchtsgutEarl Lasley

Monsanto Farmers HybridF. Douglas Lawrason

Schering Corp.Bernard Leese

Plant Variety Protection OfficeStanley Leibo

Oak Ridge National LaboratoryMorris Levin

Environmental Protection AgencyHerman Lewis

National Science Foundation

Peter LibassiVerner, Lipfert, Bernh ard & M cPherson

Paul J. Luckern

U.S. Department of JusticeClement Markert

Yale UniversityRalph R. Maurer

U.S. Meat Animal Research Center