Evidence for two genetically distinct DNA primase activities specified by plasmids of the B and I incompatibility groups.

Abstract

Plasmid ColIb-P9 of the I alpha incompatibility group is known to encode a DNA primase that acts in the conjugal transfer of the plasmid and can substitute for mutant dnaG gene product in vegetative replication of the Escherichia coli chromosome. The relevant genetic determinant (sog) has previously been cloned into a small multicopy vector plasmid. Prototype IncB plasmid R16 also suppresses host dnaG mutations. The equivalent gene(s) (pri) of R16 was cloned into plasmid pBR325 and shown by filter hybridization studies to be different from the ColI primase gene(s). The cloned fragment carrying the pri determinant encoded an enzyme which could initiate DNA synthesis in vitro on single-stranded phage M13 DNA template, but which was antigenically distinct from ColI primase. We used the cloned primase genes as probes in colony hybridization screening of strains carrying plasmids of the IncI complex and IncB group. Plasmids R64, R144 (I alpha), R621a (I gamma), RIP72, and R864a (B) contained nucleotide sequences homologous with the cloned ColI sog genes. Plasmids R805 (I xi), R724, (I delta), TP125, and PLG101 (B) showed sequence homology with the R16 pri determinant.