Copy-number mutants of the plasmid carrying the replication origin of the Escherichia coli chromosome: evidence for a control region of replication.

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RESUMO

A composite plasmid (pXX11) was constructed by joining of an oriC plasmid (pMCR115) carrying the replication origin (oriC) of the Escherichia coli chromosome and a mini-F plasmid (pSC138) carrying the ampicillin-resistance gene (bla). Plasmid pXX11 can replicate, by using oriC, in Hfr cells and mafA mutant cells that cannot support replication of an F plasmid. This plasmid is stably maintained in these host cells during cell growth even under nonselective conditions by use of the partition mechanism of the mini-F genome. In contrast to other oriC plasmids reported previously, pXX11 has no detectable effect on host cell growth. Higher copy-number (Cop-) mutants of pXX11 were isolated, and some of them were found to carry an insertion or deletion within a region derived from the E. coli chromosome. This region, designated cop (copy number), covers about 0.7 kilobase pair and is located approximately 3 kilobase pairs away from the oriC region at the side opposite the asn gene. Evidence suggests that the normal cop region locted on the oriC plasmid acts to reduce the copy number of the plasmid. Plasmid pXX11 complements the uncB402 mutation located on the host chromosome, but some of the Cop- plasmids do not, suggesting that the cop region is vey closely linked to uncB.

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