A panel of five hybrid cell lines containing mouse X chromosomes with various deletions has been obtained by fusing splenocytes from male mice carrying one of a series of reciprocal X-autosome translocations with the azaguanine-resistant Chinese hamster cell line CH3g. These hybrids have been extensively characterized by using the allozymes hypoxanthine/guanine phosphoribosyltransferase (encoded by the Hprt locus) and alpha-galactosidase (Ags) and a series of 11 X-chromosome-specific DNA probes whose localization had been previously established by linkage studies. Such studies have established the genetic breakpoints of the T(X;12)13Rl and T(X;2)14Rl X-autosome translocations on the X chromosome and provided additional information as to the X-chromosome genetic breakpoints of the T(X;16)16H, T(X;4)7Rl, and T(X;7)6Rl translocations. The data establish clearly that both the T(X;4)7Rl and T(X;12)13Rl X-chromosome breakpoints are proximal to Hprt, the breakpoint of the former being more centromeric, lying as it does in the 9-centimorgan interval between the ornithine transcarbamoylase (Otc) and DXPas7 (M2C) loci. Similarly, it is now clear that the T(X;16)16H X-autosome translocation breakpoint lies distal to the DXPas8 (St14-1) locus, narrowing the X-chromosome breakpoint down to a region flanked proximally by this marker and representing, as expected from previous data, the distal quarter of the Hprt-Ta subchromosomal span. These five hybrid cell lines provide, with the previously characterized EBS4 hybrid cell line, a nested series of seven mapping intervals distributed along the length of the mouse X chromosome. Their characterization not only allows further correlation of the genetic and cytological X-chromosome maps but also should permit the rapid identification of DNA probes specific for particular regions of the mouse X chromosome.