ISOLATION OF LAMBDA TRANSDUCING PHAGE WITH THE bio GENES INSERTED BETWEEN LAMBDA GENES P AND Q

Nyles W. Charon ¹, Allan M. Campbell ², and S. Craig Stamm ¹

¹ Department of Microbiology, West Virginia University, Morgantown, West Virginia 26506
² Department of Biological Sciences, Stanford University, Stanford, California 94305

Plaque-forming, biotin-transducing phages were constructed with the bio genes inserted between lambda genes P and Q. These phages were isolated for the eventual aim of fusing the Q gene to the bio operon. The following steps were used to construct these phages: A defective temperature-sensitive lysogen was constructed with the bio genes adjacent to and to the left of lambda genes ßNcI857OPQSRA. Heat-resistant survivors were screened for deletions with endpoints in the bio operon and to the right of P and to the left of A. Five of approximately 1,600 heat-resistant survivors had these properties. Two had the gene order bioAB.... QSRA. When these two strains were lysogenized with cI857b221 and heat induced, the desired transducing phages were obtained. We characterized these phages and studied one in detail. Two-thirds of the plaque-forming transducing phages isolated carried the entire bioB gene and only part of the bioA gene, and one-third carried the entire bioA and bioB genes. The phages isolated lost the bio genes upon propagation, indicating that they contain a partial duplication of phage genes. The duplication was shown not to involve the entire Q gene in one of these phages, bioq1b221. A recombinant of this phage, Nam7am53c17bioq1b221, failed to form plaques under biotin-derepression conditions. We conclude that if the Q gene was fused to the bio operon in this phage, not enough Q gene product was made to allow phage propagation.