bacterial genetics screencasts - introduction to conjugation

what similar processes do bacteria have to eukaryotes?

sexual reproduction and recombination

who hypothesised that bacteria could have sexual processes?

Lederburg and Tatum

what did Lederburg and tatum devise to test their hypothesis?

a selective technique to screen E. coli

what strains did Lederburg and Tatum manufacture for their experiment?

• 2 different multiply-mutant strains

• strain A = required methionine and biotin

• strain B = required threonine, leucine and thiamine

how did Lederburg and Tatum select the strains they used?

selection of mutations that had autotrophic markers (specific nutritional requirements)

what are genetic markers?

nutritional requirements and other phenotypes (e.g. resistance) - alleles used to identify strains

what did Lederburg and Tatum do with strains And B?

mixed them, incubated them and spread them on a plate containing unsupplemented medium

what did they find after growing the strains on unsupplemented medium?

some of the bacteria had regained the ability to grow on unsupplemented medium without any help

what did the bacteria that had regained the ability to grow on unsupplemented medium have?

a recombination of markers found in the parents

what happened when the strains were plating without being mixed together?

no colonies arose

what did the fact that no colonies arose when the strains were plated separately show?

reverse mutations could not occur with enough frequency for each colony to grow alone on the plate

what is a reverse mutation?

when a mutant organism produces a wild-type gene

what was suggested had taken place to allow the bacteria to grow on unsupplemented medium again?

some form of recombination

what did the U-tube experiment from Davis show?

• physical contact between the two strains was needed for genetic exchange - i.e. conjugation

• disproved that cells were leaking substances that the other cells could use for growth

how did the U-tube experiment work?

• strains placed in medium either side of a filter

• pores of the filter too small for bacteria to pass through but allowed liquid medium with dissolved contents

• none of the bacteria grew in the unsupplemented medium as they had no contact

what did one of the strains have which was the ‘donor’?

DNA molecule F (fertility plasmid)

did both strains have F (fertility plasmid)?

no, one didnt have it so was the recipient (F minus)

where and how can plasmids replicate?

• autonomously (on their own)

• not in the chromosome

what must plasmids code for to be transmitted?

the transfer (conjugation) process

how is F transmitted vertically?

via asexual divisions from mother to daughter cell

how is the F plasmid transmitted horizontally?

from cell to cell contact

what does the F plasmid induce?

pili synthesis - a cytoplasmic bridge (pore) forms between the cells

what happens when the F plasmid is transmitted?

• one strand of F is given

• DNA replication restores the other strand in the donor and the recipient

what does it mean if a strain is F+?

it has the F plasmid and is a donor

what does it mean if a strand is F-?

it does not contain the F plasmid and is a recipient

where were the genes from in Lederburg and Tatum’s experiment?

from the chromosome

what is important about the genes from lederburg and tatum’s experiment being located in the chromosome?

• they were not on the F plasmid which is the bit that can be transmitted

• a process must have occurred to get the genes onto an F plasmid

how were the chromosomal genes in the experiment tranferred?

• it was mediated by cells with F integrated into the chromosome (HFR)

• the F plasmid can be incorporated into the chromosome via homologous recombination

• HFR can still direct pili synthesis

• F promotes transfer of chromosomal DNA in conjugation

• which sets up an opportunity for recombination in the recipient

• combining the donor fragment with its own chromosome

• between the sites of recombination

what does HFR stand for?

High Frequency Recombination

what is special about the position and number of recombination events in the recipient chromosome

it is random

what does the location of the crossovers determine in the recipient chromosome?

the combination of genetic markers generated in the recipient

where and in what orientation can F integrate in the E. coli chromosome?

• at multiple positions

• in one of two orientations