DNA has been isolated from 3 human basal cell carcinomas, as well as several tumor cell lines carrying known oncogenes. These genomic DNAs were restriction-digested, electrophoresed, and probed with oligonucleotide probes to detect single-base mismatches in the Harvey, Kirsten, and N-ras proto-oncogenes at the codon 12 region and codon 61 region. Thus far, one of the skin carcinomas has been found to carry a mutant Ki-12-ras allele. We transfected the UV-irradiated cloned c-Ha-ras gene and the c- Ki-ras minigene into NIH 3T3 cells to locate sites of activation. Transformed foci were recovered. A number of foci were also obtained with unirradiated plasmids; we believe this to be due to methylation of cytosines at active sites during propagation of the plasmid in E. coli, followed by deamination. Therefore, we are repeating the experiment after subcloning all proto-oncogenes into the same vector and transforming into a non-methylating (dcm-) host.
