The expression of Streptomyces and Escherichia coli drug-resistance determinants cloned into the Streptomyces phage phi C31.
Lysogens obtained by infecting Streptomyces albus G with a phi C31-pBR322 chimaeric prophage or its delta W12 deletion derivative had increased tetracycline resistance. The ability of the delta W12 derivative to transduce tetracycline resistance was inactivated by inserting a viomycin resistance determinant (vph) into the BamHI site of the pBR322 tet gene, and restored by excising the vph gene. Another deletion mutant (delta W17) of the chimaera, carrying an intact tet gene, was normally unable to transduce tetracycline resistance. This inability was correlated with the finding, by Southern hybridisation analysis, that the att site required for insertion of phi C31 prophage into the host chromosome was located within the delta W17 deletion. Use of phi C31 lysogenic recipient permitted the integration of the att-deleted phage, presumably by homologous recombination, giving tetracycline-resistant double lysogens. This technique was extended to S. coelicolor A3(2) in the detection of derivatives of the att-deleted phage into which a thiostrepton-resistance determinant (tsr) had been inserted in vitro. Phage released from double lysogens were mainly recombinants. One such recombinant is a PstI vector for DNA cloning, able to accommodate up to 6 kb of introduced DNA.
D001435:Bacteriophages / Q000235:genetics*; D003001:Cloning, Molecular; D004262:DNA Restriction Enzymes; D004274:DNA, Recombinant / Q000378:metabolism*; D004352:Drug Resistance, Microbial; D004926:Escherichia coli / Q000187:drug effects / Q000235:genetics*; D005838:Genotype; D009693:Nucleic Acid Hybridization; D013045:Species Specificity; D013302:Streptomyces / Q000187:drug effects / Q000235:genetics*; D013752:Tetracycline / Q000494:pharmacology
D004274:DNA, Recombinant; D004262:DNA Restriction Enzymes; D013752:Tetracycline