The precise chromosomal localization of the type II renal-specific Na+-phosphate (Pi) cotransporter (NPT2) gene (gene symbol SLC17A2) is necessary for the identification of closely linked polymorphic markers to determine whether NPT2 is a candidate gene for inherited disorders of renal Pi reabsorption. Recent studies by two different groups localized NPT2 to human chromosome 5q35 and 5q13, respectively. To resolve this discrepancy, we used three independent methods. The results using a human chromosome 5/rodent somatic cell hybrid deletion panel, fluorescence in situ hybridization with a PAC clone containing the NPT2 locus, and analysis of a chromosome 5-specific radiation hybrid panel were all consistent with the 5q35 assignment of the NPT2 gene. The radiation hybrid results placed NPT2 between polymorphic microsatellite markers D5S498 and D5S469. These findings will allow the initiation of linkage analysis to determine if NPT2 has a causative role in Mendelian disorders of renal Pi wasting.