102 MUTATIONS 



Lederberg: Is that the only thing that comes off? 



Freese: Yes, this is the only thing that comes off. There is no adenine 

 that we can observe. The amomit of adenine must be less, or the ratio 

 of adenine to guanine coming off must be less than 1 to 50. 



Zamenhof: Adenine is also alkylated (60) . 



Freese: Yes, adenine is alkylated. But Lawley has shown that it is 

 preferentially alkylated in the 1- and the 3-position. 



Benzer: How much docs this treatment correspond to in terms of 

 the conditions used for mutagenesis? 



Freese: That's hard to judge because, first of all, we didn't do it 

 on phage DNA but on herring sperm DNA, and secondly, we used 

 diethyl sulfate, which reacts much stronger and much faster, instead 

 of ethylethane sulfate. But it is an ethylating agent. 



Auerbach: Is it mutagenic? 



Freese: Yes, diethyl sulfate is mutagenic, but it is very difficult to 

 work with as a mutagenic agent because it reacts so fast with water. 



Auerbach: Loveless got mutations. 



Freese: Yes. It does induce mutations, but it is harder to work 

 quantitatively with it. So it is difficult to compare quantitatively the 

 chemical and the mutagenic experiments. That, naturally, has to be 

 done. But I think it is clear that there is efficient depurination ; other 

 experiments by the Zamenhof group with bacteria and by ourselves 

 with phages have shown that depurination — in Zamenhof's case with 

 high temperatures (90) and in our case with low pH (23) — causes 

 mutations. It is likely that the main mutagenic effect of at least this 

 alkylating agent is due to the occasional removal of guanine from 

 DNA (31,48). 



Auerbach: Would that mean that it always produces deficiencies 

 when the purine goes off? 



Freese: This is another point. At the time we did the experiment 

 with the lower pH treatment, we didn't know what kind of changes 

 would be produced, but it turns out that most of the mutants we get 

 are quite similar to mutants induced, or behave in the same way as the 

 mutants induced, by base analogues (24) ; in other words, they are 

 presumably due to changes from AT to GC, and vice versa. 



Auerbach: How would that come about? 



Freese: It could have been that the gap prevented the DNA to 

 duplicate across this place or it could have been that the gap causes a 

 loop and a larger deletion. But the results indicate that DNA does 

 duplicate across the gap and just incorporates one of the four bases 

 opposite to the gap. Interestingly enough, the bases which seem to be 



