Signal strains that can detect certain DNA replication and membrane mutants of Escherichia coli: isolation of a new ssb allele, ssb-3.
Mutations in several dna genes of Escherichia coli, when introduced into a strain with a lac fusion in the SOS gene sulA, resulted in formation of blue colonies on plates containing 5-bromo-4-chloro-3-indolyl-beta-D-galactoside (X-Gal). Unexpectedly, several lines of evidence indicated that the blue colony color was not primarily due to induction of the SOS system but rather was due to a membrane defect, along with the replication defect, making the cell X-Gal extrasensitive (phenotypically Xgx), possibly because of enhanced permeability to X-Gal or leakage of beta-galactosidase. (i) In most cases, beta-galactosidase specific activity increased only two- to threefold. (ii) Mutations conferring tolerance to colicin E1 resulted in blue colony color with no increase in beta-galactosidase specific activity. (iii) Mutations in either the dnaA, dnaB, dnaC, dnaE, dnaG, or ssb gene, when introduced into a strain containing a bioA::lac fusion, produced a blue colony color without an increase in beta-galactosidase synthesis. These lac fusion strains can serve as signal strains to detect dna mutations as well as membrane mutations. By localized mutagenesis of the 92-min region of the chromosome of the sulA::lac signal strain and picking blue colonies, we isolated a novel ssb allele that confers the same extreme UV sensitivity as a delta recA allele, which is a considerably greater sensitivity than that conferred by the two well-studied ssb alleles, ssb-1 and ssb-113. The technique also yielded dnaB mutants; fortuitously, uvrA mutants were also found.
Schmellik-Sandage
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Department of Biological Sciences, Purdue University, West Lafayette, Indiana 47907.
Tessman
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D000483:Alleles; D010582:Bacteriophage lambda / Q000254:growth & development / Q000528:radiation effects; D004260:DNA Repair; D004261:DNA Replication; D004307:Dose-Response Relationship, Radiation; D004926:Escherichia coli / Q000235:genetics* / Q000528:radiation effects; D005798:Genes, Bacterial; D005838:Genotype; D009154:Mutation; D013014:SOS Response, Genetics; D013045:Species Specificity; D013489:Suppression, Genetic; D014466:Ultraviolet Rays