Our aim is to identify cellular genes whose transcriptional suppression by the v-src oncogene contributes directly and specifically to the transformed and tumorigenic phenotype. We used a modified PCR-based subtractive hybridization technique to isolate 9 cDNAs whose abundance in NIH3T3 fibroblasts is decreased 3-15-fold following transformation by the activated oncogene, v-src. Sequence analysis reveals that 3 cDNAs are unlike those in GenBank. The remaining 6 cDNAs are indentical or highly similar to rat helix-destabilizing protein gene (hnRNP A1), mouse CTLA-2 alpha cysteine protease, rat cytochrome c oxidase (COX) VIc subunit, mouse Type I collagen, human gravin and a partial human cDNA (clone A7C09) isolated by random automated sequencing. Northern blot analysis indicates that the basal level of transcripts in untransformed NIH3T3 of all the genes except mouse Type I collagen was at least 10-fold lower than that of HMG Co-A Reductase, which is abundantly transcribed. These data suggest that the down-regulation of some or all of these genes contributes to v-src-induced changes in mitogenic control or cell morphology.