Fig. 5-2

Plating bacteria and growing colonies

Commonly used genetic markers

• Prototrophic markers: wild-type bacteria are prototrophs(grow on minimal medium)

• Auxotrophic markers: mutants that require additional nutrient(fail to grow on minimal medium)

• Antibiotic-sensitivity: wild-type bacteria are susceptible(fail to grow on antibiotic-containing medium)

• Antibiotic-resistance: mutants that grow in presence of antibiotic(grow on antibiotic-containing medium)

Fig. 5-1

Chapter 5: Genetics of bacteria and their viruses

Gene transfer mechanisms in bacteria (especially E. coli)

Conjugation: orderly, deliberate transfer of DNA from one cell to another; programmed by specialized genes and organelles.

Transformation: uptake of environmental DNA into a cell

Transduction: transfer of DNA from one cell to another mediated by a virus

Properties of gene transfer in bacteria

• All are unidirectional (donor – recipient)

• Recombination requires two steps:

1. Transfer of DNA into the recipient cell, forming a

merozygote (various gene transfer mechanisms)

2. Crossing over that replaces a portion of the recipient genome (endogenote) with the homologous portion of the donor genome (exogenote)

• Transfer is always partial

Conjugating E. coli

pili

Fig. 5-6

Fig. 5-7

Conjugation in E. coli is based on the F (fertility) plasmid

Replication-coupled transfer of F

Fig. 5-8

F can integrate into the bacterial chromosome

Hfr: high frequency recombination derivative

Fig. 5-10

Transfer of integrated F includes donor chromosome

Crossing over of exo/endogenote results in recombinant genome

(replacement of a segment of recipient genome with the homologous segment of the donor genome)

Unidirectionaltransfer……

Recombination…..

Partial transfer…..

DNA transfer during conjugation is time-dependentTransfer of an entire E. coli donor genome requires

about 1 hour (F sequence is last to transfer)

Therefore, can map the chromosome as a time function:

• Mix donor Hfr and recipient F- cells

• Interrupt transfer of DNA at various times (violent mixing in a Waring blendor works!)

• Plate out cells to determine which genes were transferred within each timeframe

Fig. 5-11

Hfr azir tonr lac+ gal+ strs X F- azis tons lac- gal- strr

Fig. 5-11

Hfr azir tonr lac+ gal+ strs X F- azis tons lac- gal- strr

Genetic map generated by interrupted mating experiment

Conjugation map depends upon:

• site of F factor insertion within Hfr chromosome (original F insertion can occur at any one of many sites within chromosome)

• direction/orientation of the F factor within that Hfr strain (clockwise or counter-clockwise)

Mapping using different Hfr strains can provide a map of the entire bacterial chromosome

Fig. 5-13

Fig. 5-16

Mapping of small regions by recombination

Fig. 5-17

F integration by recombinationof IS element

Excision usinganother IS elementresults in F bearing

chromosome fragment (F’)

Transfer create partial diploid

…at least 10 species ancestors.

Fig. 5-18

Transformation: DNA in the environment of a cell is taken into the recipient cell forming a merozygote; then recombination occurs

• occurs naturally in some bacteria (e.g., Pneumococcus)

• occurs rarely in others, but can be promoted by treating cells to destabilize their membranes (e.g., in recombinant DNA work)

• can map genes by co-transformation (frequency with which two genes are simultaneously transferred

Fig. 5-19

Transduction: Transfer of DNA from one cell to another mediated by a virus; followed by recombination to integrate the DNA into the recipient cell

• can map genes by the frequency of co-transduction (frequency of simultaneous transfer of two genes)

Fig. 5-22

Fig. 5-23

Bacteriophage lytic cycle

Plaques (infection bursts) of bacteriophage on a lawn of E. coli

Fig. 5-24

Generalized transduction

Random DNA fragments are transferredFig. 5-27

Linkage mapping of a segment of the E. coli chromosome by co-transduction experiments with phage P1

Fig. 5-28

Fig. 5-30

Lysogenic infection: integration of a viral genome into one of many sites within the host cell chromosome where it quiescently resides

Upon specific cues, the process may be reversed, resulting in lytic infection

Fig. 5-31

Specialized transduction(genes nearest the insertion site are most efficiently transferred)

Fig. 5-

Fig. 5-

Fig. 5-

Fig. 5-