Conjugation

Bacterial Conjugation is the transfer of DNA by direct cell-to-cell contact, depends on the presence of a conjugative plasmid . Plasmids are small, double-stranded DNA molecules that can exist independently of host chromosomes. They have their own replication origins, replicate autonomously, and are stably inherited. Some plasmids are episomes, plasmids that can exist either with or without being integrated into host chromosomes.

The best-studied conjugative plasmid is F factor. It plays a major role in conjugation in E. coli, and it was the first conjugative plasmid. The F factor is about 100,000 bases long and bears genes responsible for cell attachment and plasmid transfer between specific E. coli cells. Most of the information required for plasmid transfer is located in the tra operon, which contains at least 28 genes. Many of these direct the formation of sex pili that attach the p+ cell (the donor cell containing an F plasmid) to an F- cell (figure 16.17). Other gene products aid DNA transfer. In addition, the F factor has several IS elements that assist plasmid integration into the host cell's chromosome.
Bacterial Conjugation
Thus the F factor is an episome that can exist outside the bacterial chromosome or be integrated into it nutrient medium, and then plated it on minimal medium. To reduce the chance that their results were due to a reversion or suppressor mutation, they used double and triple auxotrophs on the assumption that two or three simultaneous reversion or suppressor mutations would be extremely rare. When recombinant prototrophic colonies appeared on the minimal medium after incubation, they concluded that the two auxotrophs were able to associate and undergo recombination.

Lederberg and Tatum did not directly prove that physical contact of the cells was necessary for gene transfer. This evidence was provided several years later by Bernard Davis (1919-1994), who constructed a U-tube consisting of two pieces of curved glass tubing fused at the base to form a U shape with a glass filter between the halves. The filter allowed passage of media but not bacteria. The U-tube was filled with a growth medium and each side inoculated with a different auxotrophic strain of E. coli. During incubation, the medium was pumped back and forth through the filter to ensure medium exchange between the halves.

When the bacteria were later plated on minimal medium, Davis discovered that if the two auxotrophic strains were separated from each other by the filter, gene transfer did not take place. Therefore direct contact was required for the recombination that Lederberg and Tatum had observed.

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