smith: directed mutation 423 



in Salmonella typhimurium become established as mutant clones only 

 after two DNA replications (50). 



The importance to the problem of mutagenic specificity of these 

 recent concepts and experiments is that mutagenic activity can be 

 related to genetic molecular structure. The mapping of sites with 

 different mutation frequencies under the influence of different muta- 

 genic agents can be localized by use of Benzer's high-resolving power 

 technique (6, 7), with an accuracy that approaches the nucleotide 

 level. If it is assumed that only two kinds of nucleotide pairs are pres- 

 ent in DNA, the appearance of different "hot spots" would seem to 

 indicate that the mutability of a particular nucleotide pair is depend- 

 ent upon its position in the genome, perhaps the specific sequence in a 

 particular area of the macromolecule. 



The reversion of base analogue-induced mutants in both phage 

 (24, 25) and bacteria (37) by subsequent treatment with base ana- 

 logues is confirmatory evidence that the original mutations were 

 caused by transitions for which change in both directions would be 

 expected. Reversion by a particular mutagen of specific mutants, 

 which involve a restoration of function owing (in theory) to re-estab- 

 lishment of a nucleotide sequence, can be considered as specificity in 

 the strictest sense. This is the basis of the back-mutation test used in 

 Neurospora and bacteria and for reverting ill-type mutants in phage. 



The infrequency of base analogue inducible reversions with 

 mutants of spontaneous or proflavine origin led Freese to postulate 

 that most of these arose by a process different from "transition", name- 

 ly, "transversion", which involves a replacement of a purine by a 

 pyrimidine, or vice versa in a given DNA chain. 



Nitrous acid will produce mutations in tobacco mosaic virus 

 after in vitro treatment of isolated RNA (49, 57). Since de-amination 

 with nitrous acid converts cytosine to uracil, adenine to hypoxan- 

 thine, and guanine to xanthine (58), it is considered that these new 

 bases produced in .situ will have altered pairing properties and cause 

 the change of a nucleotide pair in subsequent replication of DNA 

 (25). The mutagenic effect would depend upon "transitions", as with 

 the base analogues. Nitrous acid has been found to be highly muta- 

 genic for E. coli (34, 35) and to produce a spectrum of mutations 

 which differs significantly in some respects from the spontaneous and 

 ultraviolet-induced spectra, but not from that caused by disintegra- 



