LOCALIZATION OF THE REPLICATION REGION OF THE PMJ101 PLASMID FROM VIBRIO-ORDALII

The 30-kb pMJ101 plasmid is found as a high-copy-number pool in all th e pathogenic strains of Vibrio ordalii examined so far. The replicatio n functions of pMJ101 were localized within a 2.4-kb EcoRV-HindIII res triction fragment by using different subclones in combination with Ba/ 31 exonuclease deletions and Tn5 insertion mutants. Recombinant clones carrying this fragment were able to replicate in Escherichia coli cel ls deficient in either DNA Polymerase I (PolA(-)) or integration host factor functions. However, the viability of recombinant plasmids conta ining the pMJ101 origin of replication was dependent on the expression of the gene encoding the DnaA protein. Electrophoretic analysis of pl asmid-encoded proteins in an in vitro transcription-translation couple d system revealed that the replication region of pMJ101 encodes a 36-k Da protein. The expression of this protein was correlated with the abi lity of different recombinant plasmids harboring this pMJ101 DNA regio n to replicate in the PolA(-) E. coli strain. Replicon typing showed t hat pMJ101 is not related to any of the plasmid incompatibility groups contained in the bank of rep probes described by M. Couturier et al. (Microbiol. Rev, 52, 375-395, 1988). (C) 1994 Academic Frets, Inc.