MUTAGENESIS 111 



Freese: Yes. We have done these experiments, not with nitrous acid 

 and not yet for ethylethane sulfonate, but for the phages treated with 

 lower pH, for which we assume that the purine goes off, so if we take 

 the case in which the guanine goes off, we would have a similar situa- 

 tion. The base analogues, bromouracil and 2-aminopurine, apparently 

 induce only changes from AT to GC and vice versa, so we could not 

 induce two of the three reversions which you mentioned. We found 

 that 70 per cent of the mutants could be induced to revert by these 

 base analogues, so they are presumably due to a transition. 



Auerbach: And could be reversed. 



Freese: Could be reverted, yes. We really have to do these experi- 

 ments now with the alkylating agents. 



Lederberg: Could I revert to a little bit of an earlier question that I 

 think was not fully answered; that is, how do you interpret this? What 

 is the maximum frequency of nonmottled plaques that you eventually 

 do get by holding your treated or your ethylethane sulfonate-treated 

 phage? That went quite high, didn't it? 



Freese: The total frequency of r and mottled plaques goes up to 1 

 per cent. The frequency of mottled over the sum of r and mottled — 



Lederberg: Goes down? 



Freese: Yes. You mean with ethylethane sulfonate? 



Lederberg: Yes. 



Freese: Without incubation it is very close to 1 ; in other words, most 

 of the plaques are mottled initially. Later, it goes down. 



Lederberg: But how far does it go down? 



Freese: I don't recall that. I wouldn't like to talk too much about 

 this because we haven't done enough on it. 



Lederberg: In fact, though, with the conditions of your experiment, 

 you get mottled plaques in two ways: either by delay in the mistake 

 in replication, or by the double-strandedness of the DNA. 



Freese: Yes, that's right. There I can refer to an experiment by 

 Green and Krieg (31), in which they treated phages with ethylethane 

 sulfonate and then made a single-burst experiment. This means they 

 infect the bacteria singly and then distribute the infected bacteria in 

 such a way that each tube receives only one infected bacterium. After 

 these bacteria have lysed they plate the content of each tube and score 

 the number of r mutants versus the number of wild-type mutants for 

 each bacterium. They have found that mutants can apparently be pro- 

 duced in every DNA duplication, with equal probability. 



Lederberg: You get mottled plaques on single burst? 



Freese: You get r and wild types, but from the ratio of the two, you 

 can conclude at what time the mutation was induced. 



