Loss of Hfr DNA from Escherichia coli merozygotes during inhibition of conjugation by nalidixic acid.

Abstract

SUMMARYThe effect of nalidixic acid on conjugal recombination was studied in matings with recipient strains carryingrecA200, a mutation which confers a thermosensitive Rec−phenotype. Addition of nalidixic acid to Hfr Nals× F−NalRrecA200matings at low temperature (35 °C) caused a sharp 10- to 20-fold decline in the yield of recombinants if plating on selective agar was delayed. Two separate processes were identified as being responsible for this decline. Those merozygotes in which the transferred DNA was free of the donor cell lost the ability to form recombinants through inactivation of this DNA, an effect which could be prevented by using exonuclease deficient (recB sbcB) recipients or by prior growth of exonuclease proficient recipients in medium containing 0·25 M sodium chloride. No more than 50% of the observed loss of recombinants could be attributed to this effect. The remaining merozygotes lost their ability for recombinant formation provided mating pairs, and presumably the displaced donor DNA strand, remained intact. This process was thought to involve withdrawal of transferred DNA (DeHaan & Gross, 1962) and was studied in isolation in matings withrecA+, orrecA200 recB sbcBrecipients. A mechanism involving re-annealing of the displaced Hfr DNA to the donor molecule as a result of nalidixic inhibition of gyrase activity in the donor causing relaxation of DNA supercoils is proposed to account for this withdrawal event.