ColE1-type vectors with fully repressible replication

Abstract

We have constructed two cloning vectors, pAM34 and pAM35, derived from pBR322, in which transcription of the replication primer RNA is under control of the lacZpo promoter/operator. These vectors contain a cloning cassette flanked by strong transcriptional terminators. They differ from each other by the presence (pAM34) or absence (pAM35) of gene lacIq. In the presence of repressor LacIq, replication is entirely dependent upon the addition of inducer. This feature allows the temporary maintenance of these plasmids, the construction of strains in which vector derivatives are stably integrated into the chromosome, and the recovery of nucleotide sequences adjacent to cloned fragments. Replication from the integrated plasmid can be adjusted to match the chromosome replication initiation rate required for cell growth in the absence of a functional origin, oriC.