Chapter 16 : Bacterial Genetics

Characteristics of Bacteria

• Lacks nucleus and membrane bound organelles

• have a single chromosome folded into a nucleoid body

• come in a variety of shapes and sizes

• some bacteria have capsules (to hide what they look like)

Bacteria have adapted to a range of habitats

• different shapes, adaptations all driven by the genetics of bacteria

• different habitats

  • land, water

  • metabolism must adapt to those enviorments

• bacteria play an essential role of natural processes (ie. nitrogen cycle)

Small fraction of bacteria are pathogens

Pathogen: bacterial strain that causes disease

• invades tissues

• may produce toxins, proteins that interfere with cell function/destroy it

• teatnus toxin → paralysis

Typical bacterial genome is composed of one circular chromsome

• usually circular

• DNA molecule condenses by supercoiling and looping

• each bacterium replicates its chromosome and then divides by binary fission (analogous to cytokinesis in eukaryotes) into two daughter cells

• tightly packed with genes

Bacterial genomes (focusing on E.coli)

• 90% of E.coli DNA encodes proteins, while 5% human DNA encodes protein

• e coli genes have no introns

Individual E.coli strains contain a subset of the E. coli pangenome

core genome: abt 1000 genes that are found in all strains

pangenome- core genome plus all genes that are found in some other strains and not others

Bacterial genomes contain transposons

Insertion sequences (IS) are like eukaryote TEs

• inverted repeats at the ends

• encodes transposase

• can disrupt gene funciton

• can rearrange bacterial genomes by causing deletions or inversions

Tn elements

composite TEs

• two nearby TEs

• flank a gene for resistance to antibiotics

• function to move between chromosomes and plasmids, often carrying cargo genes like antibiotic resistance or virulence factors.

Plasmids

• small circles of double stranded DNA (carry additional DNA)

• May contain genes that benefit host bacterium

Resistance plasmids

• can also provide resistance to antibiotics

• RPs

  • contain composite IS/Tn transposons

  • carry genes that confer resistance to multiple antibiotics

  • can be easily transferred from one bacterium to another in nature

Metagenomics

collective analysis of genomic DNA from an entire community of microbes

Metagenomes of extremophiles

Proteins that work under unusual conditions, such as Taq DNA polymerase

E coli is a versitale model organism

most studied and understood bacterial species

• normally inhabits intestines of warm blooded animals

• can grow in absence of O or air

• cells divide every 20 min

• phototrophic - can grow in minimal media

Bacteria are monoploid

All mutations express their phenotypes

• Altered colony morphology

• resistance to bactericides

• Auxotrophs - unable to reproduce in minimal media

Selection

establish conditions in which only the desired mutant will grow

Genetic screen

examine each colony for a particular phenotype

Gene transfer in bacteria an overview

Vertical gene transfer in bacteria

occurs in sexually reproducing organisms - traits are transferred from parent to offspring

Horizontal gene transfer in bacteria

traits are introduced from (potentially) unrelated individuals or from diff species

Transformation

competent cells take up DNA from surrounding environment

Natural transformation: when bacteria take up DNA fragments spontaneously from their surroundings

Artificial transformation: accomplished in the lab by making the cells competent

Natural transformation of B.subtilis

Conjugation

mating of bacteria

Demonstration of gene transfer by conjugation

Experiment found that if they mixed cells together that had different capabilities, could transfer capabilities btwn them

F plasmid contains genes required for the transfer of DNA

Donors for conjugation are F+ (carry an F plasmid)

recipients for conjugation are F- (don’t carry an F plasmid)

Process of conjugation

Formation of an Hfr chromosome (chromosome insertion)

F plasmid have 3 IS elements, which are identical to IS elements found at various positions on the bacterial chromosome

High frequency recombinant (Hfr) cells are formed when an F plasmid integrates into the bacterial chromosome through recombination btwn IS elements

Different Hfr Chromsomes

• when an f plasmid integrates into genome = episome

• Hfr strains differ in the location and orientation of integrated episomes

• Hfr strains retain all F plasmid functions and can be a donor for conjugation F- strain

Gene transfer btwn Hfr donors and F- recipients

• transfer of DNA starts in F plasmid at the origin of transfer

• chromosomal genes located next to F plasmid sequences are transferred to the recipient

• transferred chromosomal DNA recombines into homologous DNA recipient

Mapping genes by gene transfer during conjugation

interrupted mating experiment - you can’t stop process wherever you want, therefore map where genes are

• genes immediately follow origin of transfer in Hfr chromosome are transferred first

• order of transfer reflects the gene order on chromosome

Formation of F’ plasmids by excision from an Hfr chromosome

• F’ plasmids is formed by excision of F plasmid plus some adjacent bacterial chromosomal DNA

• F’ plasmids replicate independently in bacterial cells

F’ plasmid transfer

F’ plasmids can be transferred to F- cells by conjugation

• as long as plasmid remains in cell, and it has functionals ORI cell itself will not be affected no matter what DNA was taken into plasmid

F’ plasmids for complementation studies

• conjugation with F’ plasmids can make partial diploids called merodiploids

Bacterial conjugation

Refer to notes

What occurs during plasmid excision in Hfr cells?

1) removal of plasmid from bacterial chromosome

2) joining of the plasmid to the bacterial chromosome

3) breakdown of plasmid

4) entry of plasmid into bacterial cell

5) exit of plasmid from bacterial cell

In transduction, phage transfers DNA from a donor bacterium to a recipient bacterium

Bacteriophages (phages): viruses that infect, multiply, and kill various species of bacteria

Transduction: process by which phage transfers DNA from one host cell to another host cell

Virulent phages: always enter lytic cell after infecting cell, multiplying rapidly, and kill cell

Temperate phages: can enter either lytic cycle or enter an alternative lysogenic cycle

Lytic cycle - where virulent phages go

results in cell lysis and release of progeny phages

• phage injects its DNA into bacterial cell

• phage proteins are expressed and take over protein synthesis and DNA replication machinery of infected cell

• Phage DNA replication occurs

• Phage particles are assembled with phage DNA and phage protein

• infected cell bursts (lyses) → release new viral particles (lysate = pop of phage particles release)

Generalized transduction

• incorporation of random fragments of bacterial DNA form donor into bacteriophage particles

• DNA from donor cell injected into infected recipient cell

• Transduced chromosomal DNA recombines into homologous DNA recipient

Temperate phages

choice of lytic or lysogenic cell depends on many factors

• lysogen - generates lysogen cell- bacteria that harbor integrated temperate phage

• prophage- bacteria itself is harboring, lytic behavior of phage- temp phage that has integrated into host chormosome

Lytic and lysogenic modes of reproduction

Prophages:

do not reproduce the viral proteins needed for more virus particles

Lysogens can be induced to enter lytic cycle

• prophage excises from chromosome, undergo replication, form new virus particles

Integration of the phage DNA initiates the lysogenic cycle

Recombination btwnn att sites on phage lambda and the bacterial chromsome allows integration of the prophage

Excision of a prophage from a lysogen

Abnormal excision produces a specialized transducing phage

• bacterial DNA adjacent to integration site can be packaged and then transferred to a recipient cell

Identifying a mutant bacterial gene by plasmid library transformation

Penicillin interferes with synthesis of bacterial cell wall

penicillin binds to transpeptidates, inhibits its enzymatic activity and prevents cross linking to make the cell wall

The penicillin resistance gene

Pencillinase can degrade penicillin

Two main ways to create resistance

• target mutates

• something that removes inactivator