ADAPTATION TO PANTOTHENATE DEFICIENCY 22-28 



RANDOMNESS OF MUTATION IN THE 

 ABSENCE OF PANTOTHENATE 



A series of experiments were carried out to determine whether 

 or not the "secondary -adaptation" mutations occur at random in the 

 absence of pantothenate — that is, whether there is wide variation 

 in the number of mutants in a group of similar cultures. However, 

 in the absence of pantothenate, selection favors the synthesizing 

 mutants so that the variation due to difference in time of occur- 

 rence of mutation is magnified. Therefore, it is not possible to cal- 

 culate the variance, and it is possible to calculate the mutation rate 

 only by means of the Luria and Delbruck (1943) method which de- 

 pends on the number of cultures not containing mutations among a 

 series of similar cultures. 



A few hundred cells from separate colonies of culture in, isolat- 

 ed from agar containing pantothenate were inoculated into small 

 tubes containing 5 ml. of pantothenate -free medium and were allow- 

 ed to grow for about a month (table 22-6). At this time appreciable 

 growth had occurred and the entire cultures were plated out on test 

 agar plates to determine the number of mutants in each (experiments 

 in, rv, V, and VI). (In experiments I and II, a loop of inoculum was 

 taken directly from test tubes of culture lU during the "primary" 

 adaptation. The loop of cells was washed and diluted in 10 ml. of 

 sterile distilled water and a loop of these washed cells was used to 

 inoculate each of the 5 ml. of pantothenate -free broth in test tubes.) 

 These cultures were allowed to grow to near completion of growth 

 and then the entire cultures were plated on test agar plates. Since, 

 in each case, the inoculum used consisted of cells which had under- 

 gone a considerable number of cell divisions in the selective envir- 

 onment of the test medium, a possibility of variation in the inoculum 

 of the small tubes, which were plated out was introduced. This error 

 does not seem to be very great, however, because the cultures grew 

 quite uniformly and did not show very great differences in cell pop- 

 ulations after growing in the pantothenate -free medium for 4 weeks 

 (table 22-6). The number of mutants obtained from the various 

 groups of cultures is given in the table. 



From the great variation in the number of mutant colonies it 

 appears that the mutations occur at random. From the variation in 

 the sizes of the colonies it seems that growth rates of the mutants 

 are not all the same. 



