A rapid method for cloning mutagenic DNA repair genes: isolation of umu-complementing genes from multidrug resistance plasmids R391, R446b, and R471a.

摘要

Genetic and physiological experiments have demonstrated that the products of the umu-like operon are directly required for mutagenic DNA repair in enterobacteria. To date, five such operons have been cloned and studied at the molecular level. Given the apparent wide occurrence of these mutagenic DNA repair genes in enterobacteria, it seems likely that related genes will be identified in other bacterial species and perhaps even in higher organisms. We are interested in identifying such genes. However, standard methods based on either DNA or protein cross-hybridization are laborious and, given the overall homology between previously identified members of this family (41 to 83% at the protein level), would probably have limited success. To facilitate the rapid identification of more diverse umu-like genes, we have constructed two Escherichia coli strains that allow us to identify umu-like genes after phenotypic complementation assays. With these two strains, we have cloned novel umu-like genes from three R plasmids, the IncJ plasmid R391 and two IncL/M plasmids, R446b and R471a.