112 MUTATIONS 



Lederberg: Yes, but there should also be a fraction of mottled 

 plaques, which would be occurring in one of the existing generations, 

 you see, at the time of plating. It should be a substantial fraction. 



Freese: Since you got 200 phages out, that is not very substantial. 

 I really don't know whether they have observed this. 



Lederberg: That's only what? Six or seven or eight generations? Do 

 I take it, though, that you're working at such levels of inactivation 

 that you can assume that much of your phage inactivation is essen- 

 tially in terms of the active strand rather than the single strand, but 

 disregarding the mottling due to separation of the two strands? 



Freese: I really don't know what the mottling is preferentially 

 caused by in experiments with EMS or EES. 



Lederberg: I'm w^orried about nonmottling. 



Freese: I would say that the nonmottling probably means that one 

 of the two DNA strands has been killed off. 



Lederberg: Would you explain why the methyl agents are less active 

 in producing phage? This was Loveless's report. 



Freese: Yes, Loveless (47) found that the methylation is less muta- 

 genic than the ethylation, and we have confirmed this using methyl- 

 methane sulfonate and ethylethane sulfonate. When one plots the fre- 

 quency of mutants per viable phage versus the number of lethal hits — • 



Auerbach: Excuse me, but I don't know what you mean by "lethal 

 hits." Inactivation? 



Freese: The term "lethal hit" is derived from the Poisson distri- 

 bution. A lethal hit is equal to the natural logarithm of the phage titer 

 before versus the phage titer after treatment. In most cases the number 

 of lethal hits increases linearly with the time of treatment but some- 

 times it doesn't; then the lethal hits are a better measure for the 

 extent of the chemical reactions than the time of treatment. 



We find that the frequency of r mutants increases with increasing 

 lethal hits about five times faster with EES than with MMS. When 

 we do the same for reverse mutations we find that the frequency of 

 reversions is also increasing much faster with ethylethane sulfonate 

 than with methylmethane sulfonate. 



Neel: What is the relative frequency of forward and back muta- 

 tions? 



Freese: About 10"^ to lO"* for highly inducible mutants. This clearly 

 shows that there is a difference between ethylethane sulfonate and 

 methylmethane sulfonate, whether we take all of the nucleotides or 

 whether we consider one particular nucleotide, as we probably do for 

 reverse mutations. 



