Previously we mapped the gene encoding a human bile acid UDP-glucuronosyltransferase (UGT2B4) to chromosome 4. Here we report the mapping of two additional human UGT2B genes to chromosome 4 utilizing the polymerase chain reaction (PCR) and a panel of human/rodent somatic cell hybrid cell lines. A yeast artificial chromosome contig containing the UGT2B4, UGT2B9, and UGT2B15 genes was isolated, and pulsed-field gel electrophoresis and PCR revealed that several members of the human UGT2B gene subfamily are clustered within a 195-kb region of the YAC contig. These data permitted a provisional ordering of the genes as UGT2B9-UGT2B4-UGT2B15. Fluorescence in situ hybridization analysis, using the YAC DNA, permitted the regional localization of this gene cluster to chromosome 4q13.