Investigation of the relationship between DNA synthesis and transfer during mating in E. coli.

The question of whether or not replication of the Hfr chromosome is an obligatory preliminary to transfer has remained controversial since 1963 Latterly, questions have been raised on the number of male DNA strainds which are transferred. It has also been suggested that DNA synthesis in the recipient may be necessary. The effect of a specific inhibitor of DNA synthesis on mating between sensitive parents was studied. It was found that nalidixic acid inhibited formation of recombinants and that this inhibition was apparently reversible. While NAL was found to have a slight inhibitory effect on pair formation, the major cause of recombinsuit loss appeared to be a reversible inhibition of chromosome transfer. Inhibition of DNA synthesis in the recipient by amino-acid starvation or NAL was found to lead to loss of recombinants. With the use of a method developed for quantitating DNA transfer, it was found that there was a corresponding drop in male DNA detectable in these mated females. Attempts to use density labels to indicate the state of replication of male DNA transferred into females were foiled by rapid integration of male DNA into the female genome. Male DNA thus appeared not in the heavy or hybrid positions on the CsCl gradient, which it was expected that unreplicated or once-replicated male DNA would occupy, but in the light band together with the female DNA. However, under conditions of thymine starvation this integration was not complete and some of the male DNA transferred was detected at a buoyant density consistent with single-stranded DNA. This has still to be confirmed. Use of a thymidine-i>hosphorylase negative recipient was made to label DNA in the male cell only with C thymine in a mating mixutre.It was found that mating caused a stimulus in the rate of DNA synthesis in the male population.