Filter-hybridization studies show that major phenobarbital and pregnenolone-16alpha-carbonitrile-inducible cytochrome P-450 mRNAs in rats were encoded by members of separate, distinct gene families. These gene families are genetically divergent from each and show no cross-hybridization, even under low-stringency conditions. Furthermore, sequences contained in the P-450PB and P-450PCN gene families map to separate chromosomes of the mouse genome. Using mouse X Chinese hamster somatic cell hybrids (EBS cell lines), all distinguishable P-450PCN sequences were found to map to chromosome 6, whereas all P-450PB sequences were located on chromosome 7. Our data support the proposition that the region of the Coh locus on chromosome 7 is the site of the cytochrome P-450PB gene family. The presence of gene families for the cytochromes P-450 occurs in many mammalian species and is likely an important part of the mechanism by which the mixed-function oxidase system is capable of recognizing and metabolizing such a wide array of endogenous and foreign compounds. Conversely, NADPH-cytochrome P-450 oxidoreductase appears to be encoded in many vertebrate species by a single gene and is located on chromosome 6 of the mouse. Corroboratory data are presented to show that the Eph-1 locus on chromosome 1 is the site of at least one microsomal epoxide hydratase gene.