HAAS, ET AL.: MUTATION INDUCTION IN BACTERIA 149 



The mutations followed were those giving aberrant colonial 

 color response on EMB agar (color mutants). Synchronized cell cul- 

 tures were incubated for 1 hour in M medium supplemented with 

 either casein hydrolysate (2 mg/ml), purines and pyrimidines (0.01 

 mg/ml each of adenine, guanine, cytosine, and uracil), or with a 

 mixture of B vitamins (1 ^g/ml of each). Following incubation, ali- 

 quots were irradiated with increasing doses of UV and then dilutions 

 were plated on EMB agar. The results indicated that cells incubated 

 with purines and pyrimidines yielded somewhat higher induced 

 mutation frequencies following UV exposure. Other supplements 

 produced no changes in the subsequent UV-induced mutation fre- 

 quency from that obtained with the unsupplemented control except 

 for the growth factors, riboflavin and p-amino benzoic acid. Further 

 investigation, using various combinations of purines and pyrimidines 

 and a UV dose giving maximum mutation frequency, demonstrated 

 that maximum increase in mutation frequency was obtained when 

 uracil, cytosine, and either adenine or guanine were present. Substi- 

 tution of thymine for uracil resulted in considerable reduction in 

 the mutation frequency obtained (7). 



Experiments were next conducted to determine the duration of 

 incubation in purines and pyrimidines necessary for attaining the 

 maximum increase in UV-induced mutation. It was found that an 

 initial incubation period of 20 minutes in purine-pyrimidine medium 

 is necessary for initiation of the increase in subsequent radiation- 

 induced mutation frequency. Following this lag a rapid increase is 

 observed and the maximum frequency is attained at 30 to 35 minutes 

 incubation. On the other hand, when cells are incubated in yeast 

 extract there is no lag, and increase in susceptibility to subsequent 

 mutation induction starts immediately with incubation. This sug- 

 gested the possibility that the lag period observed with purine- 

 pyrimidine incubation represents the time necessary for synthesizing 

 radiation-reactive substances from the purines and pyrimidines. 

 Further experiments, using ribosides (adenosine, guanosine, uridine, 

 and cytidine) supported this concept since these supplements led to 

 reduction of the lag period to less than 5 minutes (7). 



Similar experiments using X-ray suggest that a large fraction of 

 X-ray-induced mutations are mediated through similar mechanisms 

 (8). Incubation of cells in yeast extract or purine-pyrimidine medium 



