Plating bacteria and growing colonies Fig. 5-2 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) Chapter 5: Genetics of bacteria and their viruses Fig. 5-1
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
onjugation in E. coli is based on the F (fertility) plasm Fig. 5-7 Replication-coupled transfer of
F F can integrate into the bacterial chromosome Hfr: high frequency recombination
Fig. 5-8derivative Transfer of integrated F includes donor chromosome Unidirectional transfer
Recombination.. Partial transfer.. Crossing over of exo/endogenote results in recombinant
genome (replacement of a segment of recipient genome with the homologous segment of the Fig. 5-10 donor genome) DNA transfer during conjugation is time-dependent
ransfer of an entire E. coli donor genome requires about 1 hour (F sequence is last to transfer) herefore, 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 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 Fig. 5-11 Genetic map generated by interrupted mating experimen
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
Mapping of small regions by recombination Fig. 5-16 F integration by recombination
of IS element Excision using another IS element results in F bearing chromosome fragment (F)
Transfer create partial diploid Fig. 5-17 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 Bacteriophage lytic cycle Fig. 5-23
Plaques (infection bursts) of bacteriophage on a lawn of E. coli Fig. 5-24 Generalized transduction
Fig. 5-27 Random DNA fragments are transferred Linkage mapping of a segment of the E. coli chromosome by co-transduction experiments with phage P1
Fig. 5-28 Lysogenic infection: integration of a viral genome into one of many sites within the host cell chromosome where it quiescently resides
Fig. 5-30 Upon specific cues, the process may be reversed,
Specialized transduction (genes nearest the insertion site are most efficiently transferred) Fig. 5-31
Fig. 5- Fig. 5- Fig. 5-
Fig. 5-
