Identification of individual sex-factor DNA strands and their replication during conjugation in thermosensitive DNA mutants of Escherichia coli

Covalent circular sex-factor DNA has been isolated from donor and recipient cells during the conjugation of normal and temperature-sensitive DNA mutants of Escherichia coli. Single strands of sex-factor DNA were centrifuged in cesium chloride-poly(U,G) gradients to give two components that have been identified by annealing experiments as the separated complementary strands. When matings are performed with either DNA temperature-sensitive donor or recipient cells, the inhibition of vegetative DNA synthesis at the restrictive temperature does not interfere with transfer and circularization of the sex-factor DNA. If DNA is isolated from temperature-sensitive donor cells mated at the restrictive temperature, a specific stimulation of sex-factor DNA synthesis can be demonstrated. By separating the complementary strands of the sex-factor in a cesium chloride-poly (U,G) gradient, this DNA synthesis has been found to be asymmetric. The sex-factor DNA strand which is synthesized in the donor has the same polarity as the strand which is transferred to the recipient.