HOXD, a gene cluster of 9 homeobox genes of the Antennapedia class; EVX2, a homeobox gene related to Drosophila-even-skipped gene; DLX1 and DLX2, two homeobox genes related to the Drosophila distal-less gene; and TTN and NEB, the genes for the two giant molecules titin and nebulin, both involved in the sarcomere structure, have been previously mapped to human 2q31-q32 and to mouse chromosome 2. We studied their relative order in human by applying FISH to three balanced chromosome rearrangements each with a breakpoint at 2q31. Unambiguous results led us to map these genes and to orient the HOXD locus along chromosome 2 according to the following order: cen, NEB, DLX1-DLX2, EVX2, HOXD (5'-3'), TTN, tel. All of these genes are part of a syntenic region covering 5-10 cM and conserved since the divergence of humans and rodents, and thus the same loci order should be present in mouse. FISH in metaphases of approximately 500 bands localized NEB to 2q24.1-q24.2, while HOXD and TTN were localized to 2q31.