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Mouse lymphoma cells of the L5178Y TK + /-3.7.2C line were exposed to varying concentrations of the anti-schistosomal drug hycanthone methanesulfonate. The trifluorothymidine (TFT)-resistant cells fell into two classes based on colony size. Southern blot analyses were performed using NcoI-digested DNA from a number of large and small mutant colonies from each treatment group. Two different restriction fragment banding patterns were identified in these analyses, those colonies that contained the 6.4 kb NcoI restriction fragment and those that did not. A total of 471 mutant colonies were analyzed and 84.5% (398) of these colonies did not exhibit the 6.4 kb fragment. There did not appear to be a hycanthone methanesulfonate dose response effect in the number of colonies that did not contain the 6.4 kb fragment among the treated groups. In addition, 82% (154 out of 188) of spontaneous mutants did not contain the 6.4 kb fragment. The results imply that greater than 80% of all spontaneous mutations found in the mouse lymphoma assay regardless of colony size do not contain the 6.4 kb fragment and each may be considered to be a large scale mutation. In addition, greater than 80% of the hycanthone induced mutations in the mouse lymphoma assay do not contain the 6.4 kb fragment and thus may be considered to be a large scale mutation.

A collection of 81 his-3 mutants of Neurospora crassa was analyzed in assays for allelic complementation and specific revertibility. In these studies, the linearity of the complementation map of the his-3 cistron (Webber, 1965) was confirmed and mutants were classified as complementing with non-polarized or polarized complementation patterns, or non-complementing. In the assays for spontaneous or induced revertibility, 89% (71/80) of the mutants reverted either spontaneously or after treatment with the chemical mutagens N-methyl-N'-nitro-N-nitrosoguanidine, ethyl methanesulfonate, 2-methoxy-6-chloro-9-(3-[ethyl-2-chloroethyl]aminopropylamino) acridine dihydrochloride, nitrous acid or hydroxylamine. The frequency of revertible mutants among the non-polarized complementing mutants was 96% (45/47), and 79% (15/19) for the polarized complementing and 79% (11/14) for the non-complementing mutants. The results of these classical genetic assays for allelic complementation and specific revertibility suggest a correlation between complementation pattern and presumptive genetic alterations at the molecular level among his-3 mutants similar to that found with ad-3B mutants induced by nitrous acid (Malling and de Serres, 1967), ethyl methanesulfonate (Malling and de Serres, 1968), or ultraviolet (Kilbey et al., 1971).