Homeobox genes play important roles in limb development. Backfoot is a recently identified mammalian homeobox gene whose temporal and spatial expression pattern during limb development suggests that it is a key component for specifying the identify and structure of the hindlimb. Here we report the chromosomal mapping of the Backfoot locus in human (BFT) and mouse (Bft). Using single-strand conformation analysis of PCR products amplified from a panel of somatic cell hybrid lines and two radiation hybrid (RH) panels, we have physically mapped BFT to human chromosome 5, closely linked to STS markers D5S2543, D5S458, D5S1947, and D5S1995 on the Stanford G3 RH map and to AFMA057VG5 and AFM350YB1 on the Gene-Bridge 4 RH map. Linkage analysis of a mouse inter-specific backcross panel (C57BL/6J x Mus musculus spretus) has localized Bft to the central part of mouse chromosome 13. The map position of Bft is near two mouse limb mutant loci defined as dumpy and mdac.