Construction and physical mapping of plasmids containing the MetA gene of Escherichia coli K-12

Plasmids containing the metA gene of E. coli K-12 were constructed in vitro using pBR322 as the cloning vehicle and λmetA transducing phage as the source of metA DNA. EcoRI digests of pBR322 and λmetA20 were joined by ligase and plasmids carrying the metA gene were selected after transformation in a metA deletion strain. Recombinant DNA molecules contained one pBR322 fragment and one λmetA20 fragment of 12.2 kb which was present in either of two possible orientations. Plasmids constructed by BamH1 digestion of λmetA2 contained a single bacterial DNA fragment of 5.8 kb inserted in the tet gene. Insertion of the metA fragment led to loss of resistance to tetracycline in one orientation and partial resistance in the opposite orientation.