Cloning of the replication gene P of bacteriophage lambda: effects of increased P-protein synthesis on cellular and phage DNA replication.
A restriction fragment of lambdaDNA carrying the P gene was cloned in the high copy number plasmid RSF2124. Cells harbouring this new plasmid RSF2124/lambdaE complement lambdaPam80 phage. A lac promoter-operator region (lacP), produced by EcoRI digestion of plasmid pKB252, was inserted into RSF2124/lambdaE such that induction of the lac promoter by IPTG or lactose leads to increased production of the P gene product. A high amount of P protein in E. coli cells results in a slow inhibition of bacterial DNA synthesis, suggesting that the initiation reaction is blocked by P protein. Synthesis of lambdaDNA proceeds normally under these conditions. Nonsuppressing groPA15 mutant bacteria which are unable to support the replication of wild-type lambda (lambdawt), acquire the ability to replicate lambdaPam80 phage but not lambdawt when they are transformed with a plasmid carrying the lambdaP gene. When harbouring a plasmid containing the mutant Pamber 80 gene, groPA15 mutants are able to support the replication of lambdawt phage when infected at a high multiplicity. lambdaPam80 phage does not multiply in these cells.
D003090:Coliphages / Q000235:genetics*; D004261:DNA Replication; D004262:DNA Restriction Enzymes / Q000235:genetics; D004269:DNA, Bacterial; D004274:DNA, Recombinant; D004279:DNA, Viral / Q000235:genetics*; D004926:Escherichia coli / Q000235:genetics; D005814:Genes, Viral; D007763:Lac Operon; D010957:Plasmids; D014764:Viral Proteins / Q000235:genetics*
D004269:DNA, Bacterial; D004274:DNA, Recombinant; D004279:DNA, Viral; D014764:Viral Proteins; D004262:DNA Restriction Enzymes