Foundations in

Microbiology Seventh Edition

Chapter 9

Microbial Genetics

Lecture PowerPoint to accompany

Talaro

Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.

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9.1 Genetics and Genes

Genetics – the study of heredity

The science of genetics explores:

1. Transmission of biological traits from parent

to offspring

2. Expression and variation of those traits

3. Structure and function of genetic material

4. How this material changes

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Levels of Structure and Function of

the Genome

• Genome – sum total of genetic material of a cell

(chromosomes + mitochondria/chloroplasts and/or

plasmids)

– Genome of cells – DNA

– Genome of viruses – DNA or RNA

• DNA complexed with protein constitutes the genetic

material as chromosomes

• Bacterial chromosomes are a single circular loop

• Eukaryotic chromosomes are multiple and linear

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Figure 9.2

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Chromosome is subdivided into genes, the fundamental unit of heredity responsible for a given trait

– Site on the chromosome that provides information for a certain cell function

– Segment of DNA that contains the necessary code to make a protein or RNA molecule

Three basic categories of genes:

1. Genes that code for proteins – structural genes

2. Genes that code for RNA

3. Genes that control gene expression – regulatory genes

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• All types of genes constitute the genetic

makeup – genotype

• The expression of the genotype creates

observable traits – phenotype

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Genomes Vary in Size

• Smallest virus – 4-5 genes

• E. coli – single chromosome containing

4,288 genes; 1 mm; 1,000X longer than cell

• Human cell – 46 chromosomes containing

31,000 genes; 6 feet; 180,000X longer than

cell

Figure 9.3 E. coli cell has spewed out its DNA

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DNA • Two strands twisted into a double helix

• Basic unit of DNA structure is a nucleotide

• Each nucleotide consists of 3 parts:

– A 5 carbon sugar – deoxyribose

– A phosphate group

– A nitrogenous base – adenine, guanine, thymine, cytosine

• Nucleotides covalently bond to form a sugar-phosphate linkage – the backbone

– Each sugar attaches to two phosphates –

• 5′ carbon and 3′ carbon

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DNA

• Nitrogenous bases covalently bond to the 1′ carbon of each sugar and span the center of the molecule to pair with an appropriate complementary base on the other strand

– Adenine binds to thymine with 2 hydrogen bonds

– Guanine binds to cytosine with 3 hydrogen bonds

• Antiparallel strands 3′ to 5′ and 5′ to 3′

• Each strand provides a template for the exact copying of a new strand

• Order of bases constitutes the DNA code

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Figure 9.4

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Significance of DNA Structure

1. Maintenance of code during reproduction

- Constancy of base pairing guarantees

that the code will be retained

2. Providing variety - order of bases

responsible for unique qualities of each

organism

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DNA Replication • Making an exact duplicate of the DNA involves

30 different enzymes

• Begins at an origin of replication

• Helicase unwinds and unzips the DNA double helix

• An RNA primer is synthesized at the origin of replication

• DNA polymerase III adds nucleotides in a 5′ to 3′ direction

– Leading strand – synthesized continuously in 5′ to 3′ direction

– Lagging strand – synthesized 5′ to 3′ in short segments; overall direction is 3′ to 5′

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• DNA polymerase I removes the RNA

primers and replaces them with DNA

• When replication forks meet, ligases link

the DNA fragments along the lagging strand

to complete the synthesis

• Separation of the daughter molecules is

complete

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Figure 9.5

DNA replication is semiconservative because

each chromosome ends up with one new

strand of DNA and one old strand.

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Figure 9.7 Completion of chromosome replication

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9.2 Applications of the DNA code

• Information stored on the DNA molecule is

conveyed to RNA molecules through the

process of transcription

• The information contained in the RNA

molecule is then used to produce proteins in

the process of translation

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Gene-Protein Connection

1. Each triplet of nucleotides on the RNA specifies a particular amino acid

2. A protein’s primary structure determines its shape and function

3. Proteins determine phenotype. Living things are what their proteins make them.

4. DNA is mainly a blueprint that tells the cell which kinds of proteins to make and how to make them

Figure 9.9 DNA-protein relationship

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RNAs

• Single-stranded molecule made of nucleotides

– 5 carbon sugar is ribose

– 4 nitrogen bases – adenine, uracil, guanine, cytosine

– Phosphate

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RNA • 3 types of RNA:

– Messenger RNA (mRNA) – carries DNA message

through complementary copy; message is in triplets

called codons

– Transfer RNA (tRNA) – made from DNA;

secondary structure creates loops; bottom loop

exposes a triplet of nucleotides called anticodon

which designates specificity and complements

mRNA; carries specific amino acids to ribosomes

– Ribosomal RNA (rRNA) – component of ribosomes

where protein synthesis occurs

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Figure 9.10

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Transcription:

The First Stage of Gene Expression

1. RNA polymerase binds to promoter region upstream

of the gene

2. RNA polymerase adds nucleotides complementary

to the template strand of a segment of DNA in the 5′

to 3′ direction

3. Uracil is placed as adenine’s complement

4. At termination, RNA polymerase recognizes signals

and releases the transcript 100-1,200 bases long

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Figure 9.11

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• All the elements needed to synthesize protein are brought together on the ribosomes

• The process occurs in five stages: initiation, elongation, termination, and protein folding and processing

Translation:

The Second Stage of Gene Expression

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Figure 9.12

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The Master Genetic Code

• Represented by the mRNA codons and the

amino acids they specify

• Code is universal

• Code is redundant

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Figure 9.13

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Figure 9.14

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• Ribosomes assemble on the 5′ end of an mRNA transcript

• Ribosome scans the mRNA until it reaches the start codon, usually AUG

• A tRNA molecule with the complementary anticodon and methionine amino acid enters the P site of the ribosome and binds to the mRNA

Translation

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Translation

• A second tRNA with the complementary anticodon fills the A site

• A peptide bond is formed

• The first tRNA is released and the ribosome slides down to the next codon

• Another tRNA fills the A site and a peptide bond is formed

• This process continues until a stop codon is encountered

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Translation Termination

• Termination codons – UAA, UAG, and

UGA – are codons for which there is no

corresponding tRNA

• When this codon is reached, the ribosome

falls off and the last tRNA is removed from

the polypeptide

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Figure 9.15

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Polyribosomal complex allows for the synthesis of

many protein molecules simultaneously from the

same mRNA molecule.

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Eukaryotic Transcription and

Translation

1. Do not occur simultaneously – transcription occurs in the nucleus and translation occurs in the cytoplasm

2. Eukaryotic start codon is AUG, but it does not use formyl-methionine

3. Eukaryotic mRNA encodes a single protein, unlike bacterial mRNA which encodes many

4. Eukaryotic DNA contains introns – intervening sequences of noncoding DNA – which have to be spliced out of the final mRNA transcript

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Figure 9.17

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Genetics of Animal Viruses

• Viral genome - one or more pieces of DNA

or RNA; contains only genes needed for

production of new viruses

• Requires access to host cell’s genetics and

metabolic machinery to instruct the host cell

to synthesize new viral particles

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9.3 Regulation of Protein Synthesis

and Metabolism

• Genes are regulated to be active only when

their products are required

• In prokaryotes this regulation is coordinated

by operons, a set of genes, all of which are

regulated as a single unit

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Operons

• 2 types of operons:

– Inducible – operon is turned ON by substrate:

catabolic operons - enzymes needed to metabolize

a nutrient are produced when needed

– Repressible – genes in a series are turned OFF by

the product synthesized; anabolic operon –

enzymes used to synthesize an amino acid stop

being produced when they are not needed

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Lactose Operon: Inducible Operon

Made of 3 segments:

1. Regulator – gene that codes for repressor

2. Control locus – composed of promoter and

operator

3. Structural locus – made of 3 genes each coding

for an enzyme needed to catabolize lactose – b-galactosidase – hydrolyzes lactose

permease – brings lactose across cell membrane

b-galactosidase transacetylase – uncertain function

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Lac Operon

• Normally off

– In the absence of lactose, the repressor binds with the operator locus and blocks transcription of downstream structural genes

• Lactose turns the operon on

– Binding of lactose to the repressor protein changes its shape and causes it to fall off the operator. RNA polymerase can bind to the promoter. Structural genes are transcribed.

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Figure 9.18

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Arginine Operon: Repressible

• Normally on and will be turned off when

the product of the pathway is no longer

required

• When excess arginine is present, it binds to

the repressor and changes it. Then the

repressor binds to the operator and blocks

arginine synthesis.

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Figure 9.19

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9.4 Mutations:

Changes in the Genetic Code

• A change in phenotype due to a change in genotype (nitrogen base sequence of DNA) is called a mutation

• A natural, nonmutated characteristic is known as a wild type (wild strain)

• An organism that has a mutation is a mutant strain, showing variance in morphology, nutritional characteristics, genetic control mechanisms, resistance to chemicals, etc.

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Figure 9.20

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Causes of Mutations

• Spontaneous mutations – random change

in the DNA due to errors in replication that

occur without known cause

• Induced mutations – result from exposure

to known mutagens, physical (primarily

radiation) or chemical agents that interact

with DNA in a disruptive manner

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Categories of Mutations

• Point mutation – addition, deletion, or

substitution of a few bases

• Missense mutation – causes change in a

single amino acid

• Nonsense mutation – changes a normal

codon into a stop codon

• Silent mutation – alters a base but does not

change the amino acid

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Categories of Mutations

• Back-mutation – when a mutated gene

reverses to its original base composition

• Frameshift mutation – when the reading

frame of the mRNA is altered

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Repair of Mutations

• Since mutations can be potentially fatal, the cell has several enzymatic repair mechanisms in place to find and repair damaged DNA

– DNA polymerase – proofreads nucleotides during DNA replication

– Mismatch repair – locates and repairs mismatched nitrogen bases that were not repaired by DNA polymerase

– Light repair – for UV light damage

– Excision repair – locates and repairs incorrect sequence by removing a segment of the DNA and then adding the correct nucleotides

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Figure 9.21

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The Ames Test

• Any chemical capable of mutating bacterial

DNA can similarly mutate mammalian DNA

• Agricultural, industrial, and medicinal

compounds are screened using the Ames test

• Indicator organism is a mutant strain of

Salmonella typhimurium that has lost the ability

to synthesize histidine

• This mutation is highly susceptible to back-

mutation

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Figure 9.22

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Positive and Negative Effects of

Mutations

• Mutations leading to nonfunctional proteins are

harmful, possibly fatal

• Organisms with mutations that are beneficial in

their environment can readily adapt, survive, and

reproduce – these mutations are the basis of

change in populations

• Any change that confers an advantage during

selection pressure will be retained by the

population

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9.5 DNA Recombination Events

Genetic recombination – occurs when an

organism acquires and expresses genes

that originated in another organism

3 means for genetic recombination in bacteria:

1. Conjugation

2. Transformation

3. Transduction

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Conjugation

• Conjugation – transfer of a plasmid or chromosomal fragment from a donor cell to a recipient cell via a direct connection

– Gram-negative cell donor has a fertility plasmid (F plasmid, F′ factor) that allows the synthesis of a conjugative pilus

– Recipient cell is a related species or genus without a fertility plasmid

– Donor transfers fertility plasmid to recipient through pilus

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Figure 9.23 (1)

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Figure 9.23 (2)

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Conjugation

• High-frequency recombination – donor’s

fertility plasmid has been integrated into the

bacterial chromosome

• When conjugation occurs, a portion of the

chromosome and a portion of the fertility

plasmid are transferred to the recipient

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Figure 9.23 (3)

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Transformation

• Transformation – chromosome fragments

from a lysed cell are accepted by a recipient

cell; the genetic code of the DNA fragment is

acquired by the recipient

• Donor and recipient cells can be unrelated

• Useful tool in recombinant DNA technology

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Insert figure 9.23 transformation

Figure 9.24

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Transduction

• Transduction – bacteriophage serves as a carrier of DNA from a donor cell to a recipient cell

• Two types:

– Generalized transduction – random fragments of disintegrating host DNA are picked up by the phage during assembly; any gene can be transmitted this way

– Specialized transduction – a highly specific part of the host genome is regularly incorporated into the virus

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Figure 9.25

Generalized

transduction

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Figure 9.26

Specialized

transduction

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Transposons

• Special DNA segments that have the capability of moving from one location in the genome to another – “jumping genes”

• Cause rearrangement of the genetic material

• Can move from one chromosome site to another, from a chromosome to a plasmid, or from a plasmid to a chromosome

• May be beneficial or harmful

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Figure 9.27

Transposons