Studies on the control of DNA replication in Escherichia coli.

On the assumption that the replication of plasmids is under negative control, an attempt was made to isolate temperature-sensitive mutants of an Flac+ factor having uncontrolled replication. It was anticipated that such mutants would be lethal to the host at 420°C but give a high frequency of Lac revertants due to loss of the F particle. Although several temperature-sensitive mutants were found, none were of this type. In some of the isolated mutants, the reversion frequency was enhanced by the presence of the F particle. On detailed study, one of these mutants proved to have a chromosomal lesion similar to those classified as defective in the initiation of chromosome replication. Known initiation-defective mutants (CRT83 and CRT46) also show an enhanced reversion frequency when infected with F or some F prime factors. In many of the revertants isolated from F+ and Flac+ derivatives of CRT83, the F factor is integrated into the chromosome. Furthermore, such revertants still carry the temperature-sensitive mutation, showing that the reversion is by suppression. It is supposed that in CRT83 and CRT46, the F replication is not temperature-sensitive and when the F particle is integrated into the chromosome the whole genome can replicate at non-permissive temperature under the control of the F replication system. However, the integration of the F particle into the chromosome alone is apparently not sufficient for the suppression of temperature-sensitivity. Studies of DNA synthesis of F+, Flac+ and Hfr derivatives of CRT46 and CRT83 showed that the temperature-sensitive defect of these mutants can be complemented partially by the F factor. Several mating experiments involving CRT83 and CRT46 showed that these mutants carry an additional mutation, mapping near ilv which renders the cells partially defective in recombinant production.