Five X-chromosome DNA markers were typed on 261 members of three large kindreds with Alport syndrome (hereditary glomerulonephritis). Lod scores greater than 3.0 for linkage between the disease locus and two of the markers confirmed X-linked inheritance of the disease. A decreasing gradient in the estimated recombination fractions observed when the markers were ordered on the basis of their map locations suggested that the disease locus is on the long arm distal to all the markers typed in this study. Using three-locus analysis we rejected all but three map orders for the six loci (the disease locus and five markers). In all three the Alport syndrome locus was on the long arm of the X chromosome distal to all the markers. Two types of Alport syndrome were represented in the three kindreds. Affected males in one kindred developed deafness in addition to nephritis; deafness did not occur in members of the other two kindreds. Although larger recombination-fraction estimates were obtained for all five markers in the kindreds without deafness, the difference was significant for only one marker. Evidence of heterogeneity was not found in tests using two markers. Markers distal to the disease locus are needed to determine whether two loci are responsible for the two types of Alport syndrome.