Search

We have isolated 14 new DNA markers from the human Xpter-Xp21 region distal to the Duchenne muscular dystrophy gene by targeted cloning, employing two somatic cell hybrids containing this region as their sole human material. High-resolution physical localization of these markers within this region was obtained by hybridization to two mapping panels consisting of DNA from patients carrying various translocations and deletions in distal Xp. Five markers were assigned to the pseudoautosomal region where their position on the long-range map of this region was further determined by pulsed-field gel electrophoresis. The other nine markers map to the X-specific region. Informative TaqI restriction fragment length polymorphisms were observed for four loci. One of these represents a region-specific low-copy repeated element. These 14 new markers represent useful tools for the understanding of distal Xp deletion and translocation mechanisms and for the positional cloning of disease genes in the region.

McLeod syndrome, characterized by acanthocytosis and the absence of a red-blood-cell Kell antigen (Kx), is a multisystem disorder involving a late-onset myopathy, splenomegaly, and neurological defects. The locus for this syndrome has been mapped, by deletion analysis, to a region between the loci for Duchenne muscular dystrophy (DMD) and chronic granulomatous disease (CGD). In this study, we describe a new marker, 3BH/R 0.3 (DXS 709), isolated by cloning the deletion breakpoint of a DMD patient. A long-range restriction map of Xp21, encompassing the gene loci for McLeod and CGD, was constructed, and multiple CpG islands were found clustered in a 700-kb region. Using the new marker, we have limited the McLeod syndrome critical region to 150-380-kb. Within this interval, two CpG-rich islands which may represent candidate sites for the McLeod gene were identified.

Using whole cosmids as probes, we have mapped 242 DMD/BMD deletion breakpoints located in the major deletion hot spot of the DMD gene. Of these, 113 breakpoints were mapped more precisely to individual restriction enzyme fragments in the distal 80 kb of the 170-kb intron 44. An additional 12 breakpoints are distributed over the entire region, with no significant local variation in frequency. Furthermore, deletion sizes vary and are not influenced by the positions of the breakpoints. This argues against a predominant role of one or a few specific sequences in causing frequent rearrangements. It suggests that structural characteristics or a more widespread recombinogenic sequence makes this region so susceptible to deletion. Our study revealed several RFLPs, one of which is a 300-bp insertion/deletion polymorphism. Abnormally migrating junction fragments are found in 81% of the precisely mapped deletions and are highly valuable in the diagnosis of carrier females.

Choroideremia (tapeto-choroidal dystrophy, TCD), an X chromosome-linked disorder of retina and choroid, causes progressive nightblindness and central blindness in affected males by the third to fourth decade of life. Recently, we have been able to map the TCD gene to a small region of overlap between five different, male-viable Xq21 deletions that were found in patients with TCD and other clinical features. Two families were identified in which classical, nonsyndromic TCD is associated with small interstitial deletions that are only detectable with probe p1bD5 (DXS165). To characterize these and two other deletions that were identified more recently, we have used the chromosome walking and jumping techniques to generate a set of five chromosomal-jumping clones flanking the DXS165 locus at various distances. With these clones, we could localize four of the eight deletion endpoints and the breakpoint on the X chromosome of a female with a de novo X/13 translocation and choroideremia. These studies assign the TCD gene, or part of it, to a DNA segment of only 15-20 kilobases.