106 MUTATIONS 



Dr. Freese, did you find that the degree of aeration had an influence 

 in your experiments? In one of my experiments, I kept part of the 

 stored suspension violently agitated, and this seemed to destroy most 

 of the delayed effect. 



Freese: I think here one has to be careful. We remove the ethylethane 

 sulfonate by adding thiosulfate, which reacts very strongly with the 

 ethylethane sulfonate. Some people don't remove the alkylating agent 

 after they have treated, and then you don't really know when the 

 treatment ends. 



Auerbach: Ordinary washing with centrifugation, even 5 times, did 

 not destroy the delayed effect. 



Freese: I must say that in higher organisms there is, I believe, 

 quite a strong possibility that at least many mutations are not due 

 to the direct effect on DNA, but either to the effect on nucleotides or 

 the effect on proteins. This is indicated also by the effect of the bi- 

 functional agents, which can have two effects. They can either have 

 the same one as we have regarded here, or they can cross-link two 

 molecules. 



Auerbach: No, no, there is no evidence that bifunctional agents are 

 better chromosome-breakers. I'll come to this later on. 



Zamenhof: Dr. Auerbach, did I understand you correctly that the 

 period of storage after treatment and removal of alkylating agent 

 before plating plays a role in the determination of the number of 

 mutants? 



Auerbach: Yes, for the adenine locus in Neurospora. In one 

 experiment we got 10 times as many mutants after 4 hours of storing 

 in water at 30° as in immediately plated spores. 



Lederberg: From the second result with the AT, not being reversed 

 by ethylethane sulfonate, do you then conclude that the substitution 

 of T for A is spurious, and that the base pair that has been originally 

 replaced was not an AT but was a GC? 



Freese: Yes. 



Lederberg: What is the other evidence to support that picture for 

 bromouracil? Can you tie that together? 



Freese: Well, we have done similar experiments with hydroxyla- 

 mine — 



Benzer: Excuse me, but before you leave the ethylethane sulfonate, 

 what is your prediction as to the mutants that are produced by 

 ethylethane sulfonate? 



Freese: Taking the results of depurination by low pH and high 

 temperature treatment, I would predict that of the order of at least 



