Chapter 9Genetics of Bacteria and Their Viruses:

Transformation

Conjugation

Jones and Bartlett Publishers © 2005

Types of genetic transfer in bacteria

• Transformation – does not require contact; one cell lyses, another takes up the DNA from the lysed cell.

• Conjugation – requires actual physical contact between the donor cell and the recipient.

• Transduction – involves a bacteriophage.

Phenology of cell competence (for transformation) in bacterial colonies

Overview ofBacterial Transformation

Mechanism of Bacterial Transformation

Donor DNA is trimmed and joined to host

(recipient) DNA by DNA Ligase

Cotransformation

• Cotransformation can occur for two genes near each other.

• Cotransformation is the probability of simultaneous transformation of two genes.

• If the rate of cotransformation is much higher than the product of the individual frequencies, then this implies that the two genes are close to each other in the bacterial genome.

Genetic transformation can be used to create genetic maps in bacteria

Lederberg and Tatum experiment

The sex factor of E. coli (F-plasmid) can recombine

into the circular bacterial chromosome

The fusion creates a high frequency

recombination (Hfr) strain.

Because the F factor can exist alone, or in the chromosome, it is

called an episome.

Conjugation in E. coli between a Hfr (male) and a F- (female) cell

Differences between F factor transfer and Hfr transfer

• Transfer of F factor takes 2 minutes; the entire chromosome takes 100 minutes.

• F factor is 100 kb; E. coli chromosome is 4600kb.

• Matings are usually interrupted before complete transfer of chromosomal genes (but hundreds of genes can be transferred).

Differences between F factor transfer and Hfr transfer

• Hfr cells do not convert F- recipient cells, because the final segment of F is not tranferred.

• In Hfr transfer, some of the transferred DNA fragment gets incorporated into the recipient chromosome.

• This will result in the F- cell becoming recombinant, but the Hfr cell stays the same.

Selectability

• How can recombinants be identified?

• The F- cell can have an allele that can be selected, such as antibiotic resistance.

• Recombinants for nutrient requirement (leu+) from Hfr cells can be screened.

• Selected marker (leu+)

• Counterselected marker (str-s).

• Recombination is not very efficient.

Interrupted-mating technique

The time of entry of genes into the F- cells can be used to make genetic maps

Different Hfr strains transfer genes starting at different locations and in

one of 2 different orientations (clockwise or counter-clockwise). The relative location of the genes (genetic map)

remains the same

The circular genetic map of E. coli showing some of the about 4,000 total genes

Note that the map is in minutes rather than in map units.

It takes 100 minutes to transfer the entire E. coli

chromosome to a F- cell.

Details of 2 minutes (2%) of the E. coli genetic map

The single origin of DNA

replication (oriC) in the

chromosome is located at about

84.4 minutes.

Abnormal excision of the F-factor from the Hfr chromosome can transfer chromosomal genes to the excised F-plasmid

(now named F’-factor; F’(lac) in the example shown)

F’ plasmids

Formation of a F’-Plasmid

Partial diploids or merodiploids – when F’ plasmids spread.