Bacteria : Clones and Mutation 



335 



FIGURE 36-6. Separate colonies replica-plated (right) from a master plate (left). 

 (Courtesy of N. E. Melechen.) 



(although there may be some exceptions due 

 to the failure of the velvet to place a portion 

 of the same colony on every replica plate). 

 Of course, one should also readily find re- 

 sistant bacteria on the master plate in the 

 corresponding position. Despite the fact that 

 this clone-sampling technique is advantageous 

 for many other purposes, it is still too labo- 

 rious for testing the pre- or postadaptation 

 hypothesis, since to be reasonably sure of 

 finding one clonal mutant to streptomycin- 

 resistance would require making replicas of 

 about 10 thousand plates. 



It is possible to circumvent this difficulty 

 by using a third method for clone-sampling, 

 which involves replica-plating contiguous 

 colonies. A billion or so bacteria can be 

 plated on drug-free agar. These will form 

 small clones so closely spaced as to show con- 

 tinuous growth (Figure 36-7A). Neverthe- 

 less, a replica of this growth can be made on 

 streptomycin agar. The replica will show 

 growth whenever there are drug-resistant 

 mutants (Figure 36-7B, C, D). One can then 

 return to the corresponding site on the master 

 plate and obtain samples to be tested for 

 resistance to the drug. If such a sample is 

 much richer in resistant mutants than a 



sample from a site on the master plate ran- 

 domly chosen from those not mutant on the 

 replica, the preadaptive view is proved; if 

 not, the postadaptive view is. This experi- 

 ment has been performed and the results 

 prove that most mutants are clearly pre- 

 adaptive (Figure 36-7). Moreover, other 

 experiments show conclusively, in the case 



FIGURE 36-7. Replica-plating continuous colonies 

 for the detection of mutants to streptomycin- 

 resistance. (After J. Lederberg and E. M. 

 Lederberg.) 



