Moloney murine sarcoma virus carries an oncogenic sequence (v-mos) which is homologous to a single copy gene (c-mos) present in the normal cells of several vertebrate species. Because of the possible significance of c-mos sequences in normal development and malignant transformation induced by physical or chemical agents, we have examined the state of integration, methylation, and transcriptional activity of c-mos sequences in a variety of normal rodent tissues, normal cell lines, or cell lines transformed by radiation or chemical carcinogens. DNA-DNA hybridization, utilizing the Southern blotting technique and a plasmid-derived DNA probe representing the v-mos sequence, gave no evidence for rearrangements of the c-mos sequence in the DNAs obtained from these diverse cell types. Parallel studies employing the restriction enzyme isoschizomers HpaII and MspI indicated that in all of these cell types the c-mos sequences were heavily methylated. In addition, analysis of cellular RNAs by blot hybridization with the v-mos probe failed to detect evidence of transcription of the c-mos sequences in any of these cell types. This was in contrast to a Moloney sarcoma virus-transformed cell line in which we found that the integrated v-mos sequence was both undermethylated and extensively transcribed. Thus, it would appear that c-mos sequences do not play a role in the transformation of rodent cells by chemical or physical agents, although the possible role of other endogenous onc sequences remains to be determined.