Restriction endonuclease fragment length variations (RFLVs) were found through the use of cDNA probes for metallothionein genes 1 (Mt-1) and 2 (Mt-2) in the mouse. RFLVs were detected in restriction patterns generated by BglII and XbaI in the Mt-1 gene and by PvuII in the Mt-2 gene. All laboratory strains carry the Mt-1a and Mt-2a alleles. Among strains of wild origin, some Western European subspecies (Mus mus domesticus and M. m. brevirostris) also carry the Mt-1a and Mt-2a alleles. In contrast, a European subspecies (M. m. musculus) and the great majority of subspecies from East Asian countries (M. m. molossinus, Chinese mice of wild origin, and M. m. yamashinai) carry the Mt-1b and Mt-2b alleles. A domesticus strain from Bulgaria and two castaneus strains from Thailand and Philippines carry the intermediate combination of Mt-1b and Mt-2a alleles. Using the RFLVs, we mapped the Mt-1 and Mt-2 genes on chromosome 8, and they appear to be very closely linked since no recombination was observed between them in any of the mice examined. Data from three-point cross tests showed that the recombination frequencies are 4.31% between Os and Mt, 15.52% between Mt and Prt-2, and 19.83% between Os and Prt-2. The gene order of Os-Mt-1,Mt-2-Prt-2 has been confirmed.