Hum Genet (1988) 80 :296-298 © Springer-Verlag 1988 Genetic mapping of four DNA markers (DXS16, DXS43, DXS85, and DXS143) from the p22 region of the human X chromosome Carolyn J. Brown, Melanie M. Mahtani, and Huntington F. Willard Department of Medical Genetics, Medical Sciences Building 4282, University of Toronto, Toronto, Ontario M5S IA8, Canada Summary. Linkage analysis of four polymorphic anonymous DNA markers from the Xp22 region was performed using families from the Centre d'Etude du Polymorphisme Humain. The loci DXS43 (pD2) and DXS16 (pXUT23) were found to be tightly linked (0 = 0.02 at Z = 14.96) and proximal to both DXS85 (782) and DXS143 (dic56). Multipoint linkage analy- sis suggests the order: cen-DXS43 DXS16 DXS85 DXS143-pter 2cM 3cM 13cM Introduction The Xp22 region of the human X chromosome contains se- quences distal to the Duchenne muscular dystrophy gene and proximal to the pseudoautosomal region (Davies et al. 1988). While these latter two regions have been extensively mapped, both physically and genetically, less is known regarding the linkage relationships of the region between them. This region includes a number of disease loci and several polymorphic anonymous DNA markers. We have recently derived a sec- ond polymorphic probe (Brown and Willard 1987) by a walk from the DXS16 locus (pXUT23), which had previously been localized to this region (Davies et al. 1985). We have used this probe in conjunction with three other restriction fragment length polymorphisms (RFLPs) from Xp22 in a linkage analy- sis using families from the Centre d'Etude du Polymorphisme Humain (CEPH). An estimate of the genetic distances de- lineated by these markers should be useful for linkage analysis of families affected by X-linked diseases in this region. The loci studied each has a minor allele frequency of 0.4 or greater and, therefore, should be potentially useful markers in clin- ical studies. Diseases that have been previously shown to be Offprint requests to: H. F. Willard Table 1. Summary of probes used for linkage analysis linked to one or more of the markers used in this study in- clude: hypophosphataemic rickets; ocular albinism; retino- schisis; Coffin-Lowry syndrome; and a form of X-linked men- tal retardation (Davies et al. 1988). Materials and methods Family material was obtained from the Centre d'Etude du Polymorphisme Humain (CEPH). Conditions for restriction enzyme digestion of the genomic DNAs, agarose gel electro- phoresis, transfer to nitrocellulose, prehybridization, and hy- bridization have been described (Willard et al. 1983; Mahtani and Willard 1988). The final filter wash for all experiments was carried out in 0.1 x SSC; 0.1% SDS at 65°C. Details re- garding the probes used are outlined in Table 1. Inserts iso- lated from each of the plasmids were radioisotopically labelled by random priming (Feinberg and Vogelstein 1983). Families informative at the DXS16 locus were subsequent- ly tested for polymorphisms at the other three loci. Ten of the CEPH families were informative for two or more of the mark- ers. These pedigrees were analyzed for linkage with the com- puter programs LIPED (Ott 1974) and CILINK, version 4.6 (Lathrop et al. 1986) for two-point and multilocus analysis, re- spectively. Genotypic data for these four markers have been communicated to the CEPH database. Results Figure 1 shows representative Southern blots for one of the families informative for all four markers. The lod scores and (lod -1) confidence limits (Conneally et al. 1985), derived from analysis of the complete data with the LIPED program, are detailed in Table 2. Multipoint linkage analysis favoured Locus Probe Polymorphie Allele 1 enzyme Allele 2 Reference Size Freq. Size Freq. DXS16 XUT23 -SE3.2-L MspI 7.0 -B2.1 BglII 17.5 DXS43 pD2 PvuII 6.6 DXS143 dic56 BclI 8.9 DXS85 782 EcoRI 14.0 0.53 5.5 0.47 0.84 12.5 0.16 0.55 6.0 0.45 0.56 7.4 0.44 0.60 7.0 0.40 Brown and Willard (1987) Bakker et al. (1985) Aldridge et al. (1984) Middlesworth et al, (1985) HoNer et al. (1985)