Characterization of a replication mutant of the bacteriocinogenic plasmid Clo DF13

Abstract

In a previous paper (Kool, A.J. and Nijkamp, H.J.J. (1974) J. Bacteriol. 120, 569–578) the isolation of a mutant of the bacteriocinogenic plasmid Clo DF13, Clo DF13-Rep3, has been described. It was observed that cells harbouring the mutant plasmid synthesize more plasmid DNA than cells harbouring the wild type plasmid. This paper deals with the characterization of the nature of this plasmid-specific mutation. The following properties of the Clo DF13-Rep3 mutant plasmid could be observed: 1. 1. The plasmid-specific mutation did not lead to a significant change in the sedimentation value of Clo DF13 DNA. 2. 2. The specific rate of Clo DF13-Rep3 DNA synthesis (expressed as the number of plasmid DNA molecules synthesized per min) is on average seven times the specific rate of wild type Clo DF13 DNA synthesis. 3. 3. Also chromosomeless minicells, harbouring the Clo DF13-Rep3 plasmid, contain about seven times more plasmid DNA as wild type Clo DF13-harbouring minicells. 4. 4. The replication time of the Clo DF13-Rep3 plasmid is approx. 90 s at 30°C and does not differ significantly from the replication time of the wild type plasmid. 5. 5. The Rep3 mutation did not alter the dependence of Clo DF13 plasmid replication on the dnaA and dnaC gene products. 6. 6. The plasmid-specific mutation is cis-dominant over wild type. The data presented in this paper indicate that this mutant plasmid is not affected in the elongation but in the initiation of plasmid DNA replication.