164 MUTATION AND PLANT BREEDING 



Those mutations induced in the parental DNA should require only 

 a short period of protein synthesis following induction. With muta- 

 tions to prototrophy, expression on M agar would occur as soon as 

 protein synthesis for new enzyme production takes place. Treatment 

 with CMP after this time would not interfere with their expres- 

 sion. On the other hand, mutations induced in daughter DNA, 

 presumably during the process of replication, might be dependent 

 on mutagenic precursors or on chemical mutagens produced by the 

 X-ray and acting at some stage in the implicative process. These 

 would correspond to the so-called "delayed" or "indirect" muta- 

 tions produced by X-ray. In Figure 7 these correspond to that frac- 

 tion susceptible to MFD after CMP addition at 10 minutes and 

 revealed by the broth-supplemented M agar platings. 



This hypothesis is readily testable when CMP challenge is 

 run on X-irradiated cells, and the cells then plated on both broth- 

 supplemented M agar and on M agar (Figure 8). From the supple- 

 mented M agar plates it is evident that during the first 10 minutes 

 of postirradiation incubation in complete medium about half of 

 the mutations become susceptible to CMP challenge. This sensi- 

 tivity remains constant for approximately 20 minutes and then is 

 lost with further incubation. All mutations are insensitive to CMP 

 after 30 to 40 minutes incubation. Similar results have been obtained 

 using the uracil analogue 6-AU, and are additional evidence that the 

 CMP-sensitive fraction of X-ray-induced mutants is induced in daugh- 

 ter DNA during the replicative process and that RNA and protein 

 are involved in replication. 



Other experiments designed to determine whether the CMP- 

 resistant fraction of mutants can be expressed in the absence of 

 DNA synthesis were carried out using E. coli strain 1 5 T - MoTyi .-, follow- 

 ing reversion of the tyrosine requirement in this strain. It was found 

 that immediately after irradiation few mutants were expressed on 

 M plus thymine plus methionine agar but large numbers were 

 expressed on broth-supplemented M agar. In further experiments 

 with this strain thymine was withheld from tyrosine and methionine- 

 supplemented postirradiation incubation medium for increasing 

 periods of time and then added. All platings were on thymine and 

 methionine supplemented M medium after a total postirradiation 

 incubation period of 80 minutes. In these experiments approximately 



