The construction in vitro of derivatives of bacteriophage lambda carrying the amidase genes of Pseudomonas aeruginosa

Summary The amidase genes of Pseudomonas aeruginosa were inserted into a replacement vector following cleavage with the restriction endonuclease HindIII. The recombinant ami was detected by enhanced growth of Escherichia coli around plaques of the recombinant phage on minimal medium containing acetamide as the nitrogen source. Low levels of amidase activity were detected in E. coli cultures infected with ami and these were sufficient to allow growth with acetamide as nitrogen source. Lysis-defective derivatives of ami were made by introducing Q ^-, S ^- mutations. Cultures of E. coli infected with amiQ ^- S ^- synthesised amidase as the major protein. The amidase produced by these cultures was identical to that produced by PAC strains of P. aeruginosa in substrate specificity, thermal stability and immunological crossreaction.