Cir

thr+ leu+ thi+ bio phe cys thr leu thi bio phe cys Prototrophic strain thr+ leu+ thi+ bio+ phe+ cys+

What they did not know was how it had taken place.

To study this problem, Bernard Davis constructed a U-shaped tube ( FIGURE 8.11) that was divided into two compartments by a filter having fine pores. This filter allowed liquid medium to pass from one side of the tube to the other, but the pores of the filter were too small to allow passage of bacteria. Two auxotrophic strains of bacteria were placed on opposite sides of the filter, and suction was applied alternately to the ends of the U-tube, causing the medium to flow back and forth between the two compartments. Despite hours of incubation in the U-tube, bacteria plated out on minimal medium did not grow; there had been no genetic exchange between the strains. The exchange of bacterial genes clearly required direct contact between the bacterial cells. This type of genetic exchange entailing cell-to-cell contact in bacteria is called conjugation.

F+ and cells In most bacteria, conjugation depends on a fertility (F) factor that is present in the donor cell and absent in the recipient cell. Cells that contain F are referred to as F+, and cells lacking F are F".

The F factor contains an origin of replication and a number of genes required for conjugation (see Figure 8.8). For example, some of these genes encode sex pili (singular, pilus), slender extensions of the cell membrane. A cell containing F produces the sex pili, which makes contact with a receptor on an F~ cell ( FIGURE 8.12) and pulls the two cells together. DNA is then transferred from the F+ cell to the F~ cell. Conjugation can take place only between a cell that possesses F and a cell that lacks F.

Question: How did the genetic exchange seen in Lederberg and Tatum's experiment take place?

Auxotrophic strain A

X"

Auxotrophic strain B

Airflow

-Strain B

Minimal medium

Minimal medium

8.11 Davis's U-tube experiment.

In most cases, the only genes transferred during conjugation between an F+ and F~ cell are those on the F factor ( FIGURE 8.13a and b). Transfer is initiated when one of the DNA strands on the F factor is nicked at an origin (oriT). One end of the nicked DNA separates from the circle and passes into the recipient cell ( FIGURE 8.13c). Replication takes place on the nicked strand, proceeding around the circular plasmid and replacing the transferred strand ( FIGURE 8.13d). Because the plasmid in the F+ cell is always nicked at the oriT site, this site always enters the recipient cell first, followed by the rest of the plasmid. Thus, the transfer of genetic material has a defined direction. Once inside the recipient cell, the single strand

No growth No growth No growth No growth

Conclusion: Genetic exchange requires direct contact between bacterial cells.

8.12 A sex pilus connects F+ and F~ cells during bacterial conjugation. (Dr. Dennis Kunkel/Phototake.)

(donor (recipient bacterium) bacterium)

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