biotechnology a. genetic engineering, also known as recombinant dna technology, means altering the...

BIOTECHNOLOGY

A. Genetic engineering, also known as recombinant DNA technology, means altering the genes in a living organism to produce a Genetically Modified Organism (GMO) with a new genotype.

B. Various kinds of genetic modification are possible: 1.inserting a foreign gene from

one species into another, forming a transgenic organism

2.altering an existing gene so that its product is changed

3.changing gene expression so that it is translated more often or not at all.

Humans are made up of trillions of cells

Each cell:

•46 human chromosomes

•2 metres of DNA

•3 billion DNA subunits (A,T,G,C)

•Approximately 22,000 genes

DNA RNA Proteintranscript translation

replication

genotype phenotype

A. It allows genes from one organism to be inserted into a cell of a different organism of a different species.

• Examples:– Human genes can be inserted into a

bacterium– Human genes can be inserted into cells

from other animals– Bacterium genes can be inserted into

plant cells

1. Isolation-Isolate the gene2. Restriction- Cutting3. Transformation - Insert it in a host using

a vector and produce as many copies of the host as possible

4. Expression-Separate and purify the product of the gene

E. coli bacterium

Bacterialchromosome

A plasmidis isolated.

Gene ofinterest

The plasmid is cutwith an enzyme.

Plasmid

The cell’s DNAis isolated.

The cell’s DNA is cutwith the same enzyme.

DNA

Examples of gene use

A cell with DNAcontaining the geneof interest

Geneof interest

The targeted fragmentand plasmid DNAare combined.

DNA ligase is added,which joins the twoDNA molecules.

Geneof interest

Genes may be insertedinto other organisms.

The recombinant plasmidis taken up by a bacteriumthrough transformation.

Examples of protein use

Harvestedproteinsmay beuseddirectly.

The bacteriumreproduces.

Cloneof cells

Recombinantbacterium

RecombinantDNAplasmid

1

3

5

4

2

6

7

9

8

E. colibacterium

Bacterialchromosome

A plasmidis isolated.

Gene ofinterest

Plasmid

The cell’s DNAis isolated.

DNA

A cell with DNAcontaining the geneof interest

12

E. colibacterium

Bacterialchromosome

A plasmidis isolated.

Gene ofinterest

Plasmid

The cell’s DNAis isolated.

DNA

A cell with DNAcontaining the geneof interest

1

3

2

4

The plasmid is cutwith an enzyme.

The cell’s DNA is cutwith the same enzyme.

Geneof interest

E. colibacterium

Bacterialchromosome

A plasmidis isolated.

Gene ofinterest

Plasmid

The cell’s DNAis isolated.

DNA

A cell with DNAcontaining the geneof interest

1

3

2

4

5

The plasmid is cutwith an enzyme.

The cell’s DNA is cutwith the same enzyme.

Geneof interest

The targeted fragmentand plasmid DNAare combined.

E. colibacterium

Bacterialchromosome

A plasmidis isolated.

Gene ofinterest

Plasmid

The cell’s DNAis isolated.

DNA

A cell with DNAcontaining the geneof interest

1

3

2

4

5

6

The plasmid is cutwith an enzyme.

The cell’s DNA is cutwith the same enzyme.

Geneof interest

The targeted fragmentand plasmid DNAare combined.

DNA ligase is added,which joins the twoDNA molecules.

Geneof interest

RecombinantDNAplasmid

Geneof interest

The recombinant plasmidis taken up by a bacteriumthrough transformation.

Recombinantbacterium

RecombinantDNAplasmid

7

Geneof interest

The recombinant plasmidis taken up by a bacteriumthrough transformation.

The bacteriumreproduces.

Cloneof cells

Recombinantbacterium

RecombinantDNAplasmid

7

8

Geneof interest

The recombinant plasmidis taken up by a bacteriumthrough transformation.

Harvestedproteinsmay beuseddirectly.

The bacteriumreproduces.

Cloneof cells

Recombinantbacterium

RecombinantDNAplasmid

Genes may be insertedinto other organisms.

9

7

8

Donor DNA

Genetic probe

(a) Isolation of a specific gene from donor e.g. human

• Cells broken open

• Genetic probe added

• Reveals position of the gene of interest

Position of gene of interest

Bacterial cell

Plasmid

(b) Isolation of plasmid from a bacterial cell

www.sci.sdsu.edu

Restriction enzymes (also called endonuclease) act as molecular scissors and cut DNA at specific sites called restriction sites

This makes a staggered cut with sticky ends ex: EcoRI

Restriction site

Restriction site

Restriction ezymes

Clipartguide.com

Restriction sites

Plasmid

Restriction site Restriction siteDonor DNA

Restriction enzymes

Donor DNA

Sticky EndsPlasmid

Enzyme DNA ligase bonds sticky ends together

Donor DNA with

sticky ends

Plasmid DNA with sticky ends

A. Vector – molecule of DNA which is used to carry a foreign gene into a host cell◦ Most common vectors are bacteria and viruses

B. A vector has to have certain properties: 1. It is big enough to hold the gene we

want 2. It is circular (or more accurately a

closed loop), so that it is less likely to be broken down

3. It contains control sequences, such as a transcription promoter, so that the gene will be replicated or expressed.

4. It contain marker genes, so that cells containing the vector can be identified

Plasmid DNA

Donor DNA

Enzyme DNA Ligase bonds sticky ends together

Next slide for animation

www.gch.ulaval.ca

Recombinant DNA

D. Recombinant DNA introduced into bacterial cell

Bacterial chromosome

Bacterial cell

Recombinant DNA

Bacterial cell reproduces by Binary Fisson

• Bacterial cell produces the polypeptide

• Coded for by the donor DNA

Donor DNA

Plasmid

1. Cut with restriction enzymes

Donor DNA

Sticky Ends

2. Ligase bonds sticky ends together

Recombinant DNA

www.encarta.msn.com

Genetically Engineered

Products

Insulin regulates the blood sugar level.

The gene for human insulin is inserted into yeast or bacteria, from which large quantities of the human insulin are manufactured

Production of humulin

https://www2.vaxserve.com/ index.cfm?event=get...

Undernourished people in poorer countries may have blindness caused by a lack of vitamin A

Contains beta-carotene, which forms Vitamin A

http://www.healingdaily.com/detoxification-diet/rBGH.jpg

rBGH is a genetically engineered version of a hormone, which is found in the pituitary gland of cows and controls milk production

rBGH can increase cows’ milk production by as much as 20-30%

http://simple.wikipedia.org/wiki/Image:Milk_glass.jpg

Elastic, light weight fiber 5 times stronger than steel

One method involves inserting the gene from a weaving spider into a fertilized goat egg. The resultant “spider-goat” produces milk that can be manufactured into strong fibers.

http://www.chm.bris.ac.uk/motm/spider/goat.jpg

To make flak jackets, rope, textiles, sutures, artificial tendons, and bandages for burn victims

Genes that make jellyfish glow are inserted into other organisms

The organism glows under UV light

Bt is a bacterial toxin that is toxic to some insects. In the lab, the gene that produces the toxic effect is inserted into the DNA of plants

Bt crops produce an insecticide protein thousands of times more powerful than the chemical spray insecticides

Can resist infection by an insect European corn borer

Genetically modified corn

www.farmbureaulaporteco.org

Transgenic plants and farm animalspest-resistant (reduce use of pesticides),increase yield

Increase storage time e.g. green tomato

tomato with beef genes

Clonal propagation: a source of tissue or organ for transplantationavoid all problems of immunoincompatibility.

Gene therapy:Artificially replace the disease-causing gene with a normal allele.The normal allele can be carried by a virus vector to the target tissues.e.g. treatment of cystic fibrosis

Production of pharmaceuticals for treatment of diseases e.g. human insulin, interferons

Production of pharmaceuticals for disease prevention e.g. vaccine (hepatitis B vaccine)

Use of GM microorganisms to make stone-wash jeans

Use of GM microorganisms to produce enzymes e.g. detergents

GM E. coli possesses gene to break down cellulose, speeding up recycling of the most abundant biomass on earth

GM microorganisms with enhanced ability to break down environmental pollutants

Dangerous pathogens formed in the course

New tools for militarists and terrorists Triggering of catastrophic ecological

imbalance Moral problems in the use of GM

techniques in man e.g. germ cell gene therapy

Unknown effect of GM food on men

Main concerns are in Genetically Modified Foods.

Effects on Food Chains of genetically modified plants and animals.

Also conflicts in relation to human cloning and stem cell research.