A determination of mutagen specificity in bacteria using nonsense mutants of bacteriophage T4

Abstract

We present a method for the determination of mutagen specificity in bacteria, using an Escherichia coli strain that is mutant because of an amber triplet in a gene necessary for arginine biosynthesis. A large number of revertants occurring spontaneously and after treatment with 2-aminopurine, [5- 3H]uracil radioactive decay, ethyl methanesulfonate, 5-bromodeoxyuridine, hydroxylamine and ultraviolet light were tested for their ability to support the growth of 24 amber mutants and one ochre mutant of bacteriophage T4. Regardless of the mutagen used to produce the revertants, only 6 patterns, called classes, of phage growth were obtained. Revertants of classes 1, 2 and 3 contained amber suppressors, revertants of classes 4 and 5 contained ochre suppressors and those of class 6, since they suppressed none of the mutant phages used, were assumed to be structural gene revertants. By comparing the patterns of phage growth obtained to those for bacterial strains with characterized suppressors, the amino acid inserted by a revertant of class 1, 2 or 3 was inferred. Strikingly different distributions into the classes were observed for some of the mutagens. 2-Aminopurine, ethyl methanesulfonate, and [5- 3H]uracil radioactive decay were found to be very specific in their action. Assuming that reversion occurred by single base changes in DNA specifying either the amber codon or certain sRNA anticodons, it was also possible to infer the base changes produced by the mutagens. The sRNA anticodons assumed to be altered are those that can become the inverse complements of either UAG or UAA by single base changes.