DNA FINGERPRINT ANALYSIS FOR THE DETECTION OF INDUCED MUTATIONS IN MAMMALIAN-CELLS IN CULTURE

A mutation assay in cultured mammalian cells based on the direct analy sis of minisatellite DNA was developed. Band pattern variations reflec t DNA alterations ranging from single base changes to complex rearrang ements. By DNA fingerprinting a large number of autosomal loci through out the human genome can be simultaneously checked, therefore minimizi ng the size of the samples of cell colonies to be scored in the absenc e of phenotypic selection. For the mutation assay chinese hamster cell s (V79) were treated with Nitrosoguanidine and 14 independent colonies were isolated and expanded. DNA fingerprints were obtained after dige stion of the DNA extracted from each clone with both HinfI and Hae III , and hybridisation with both 33.15 and 33.6 probes. Twelve colonies f rom untreated cells were also analysed. Several differences in the ban d pattern of treated colonies were observed when compared with untreat ed cells; digestion with Hae III and hybridisation with 33.15 probe al lowed the detection of the highest frequency of induced variants. The results suggest that minisatellite sequences are hypermutable sites th at can be used to monitor the mutagenic potential of chemical agents d irectly at the DNA level, without phenotypic selection. Moreover, with the method herein decribed, it is possible to distinguish between tru e mutations and epimutations, such as those caused by changes in DNA m ethylation.