The homeotic genes of Drosophila, which regulate pattern formation during larval development, contain a 180-base-pair DNA sequence termed the "homeo-box." Nucleotide sequence comparisons indicate that the homeo-box motif is highly conserved in a variety of motazoan species. As in Drosophila, homeo-box sequences of mammalian species are expressed in a temporal and tissue-specific pattern during embryogenesis. These observations suggest functional homologies between dipteran and mammalian homeo-box gene products. To identify possible relationships between homeo-box genes of mice and humans, we have compared the chromosomal location of homeo-box genes in these species. Using in situ hybridization and somatic cell genetic techniques, we have mapped the chromosome 6-specific murine Hox-1 homolog to the region p14-p21 on human chromosome 7. We have also regionally mapped the murine Hox-3 locus to 15F1-3 and its human cognate to 12q11-q21. These comparative mapping data indicate that a syntenic relationship in mice and humans is maintained for all homeo-box loci examined to date. We suggest these regions represent evolutionarily conserved genomic domains encoding homologous protein products that function in regulating patterns of mammalian development.