We have cloned and sequenced a cDNA encoding the mammalian homologue of Distal-less 3 (Dlx-3) from a rat embryonic brain cDNA library. The primary structure of rat Dlx-3 showed a cDNA with an open reading frame of 290 amino acids with a molecular weight of 31 kDa harboring a homeodomain sequence characteristic of the Distal-less homoeobox gene. In addition to the highly conserved homeodomain sequence, we newly found the consensus motifs in the N-terminal and C-terminal region, which were specifically conserved among Distal-less members of different species. Phylogenetic analysis of the Distal-less homeodomain sequence also showed a possible association between the sequence similarities in the homeodomain and the spatial specifications of homeobox genes expressed in the developing central nervous system.