We are exploring the origin and function of amplified DNA sequences associated with double minutes (DMs) in a spontaneously transformed derivative of mouse 3T3 cells. Toward that goal, we have constructed a cDNA library using RNA from these cells and have isolated cDNA clones representing sequences that are amplified and overexpressed in these 3T3-DM cells. From results of Northern- and Southern-blot analyses, we conclude that these cDNAs represent two distinct genes, which we have designated mdm-1 and mdm-2. Using DNAs from a panel of Chinese hamster-mouse somatic cell hybrids together with in situ hybridization protocols for gene mapping studies, we have found that these DM-associated, amplified DNA sequences originate from mouse chromosome 10, region C1-C3. Sequences homologous to mdm-1 and mdm-2 are present in the genomes of several species examined, including that of man.