ISOLATION OF A YEAST ARTIFICIAL CHROMOSOME CONTIG SPANNING THE X-CHROMOSOMAL TRANSLOCATION BREAKPOINT IN A PATIENT WITH RETT-SYNDROME

Rett syndrome is a neurodevelopmental disorder observed exclusively in females. A de novo X;3 translocation was detected in a patient (TH) w ith Rett syndrome. The X chromosomal breakpoint maps to Xp21.3 between the distal end of the Duchenne muscular dystrophy (DMD) gene and the DXS28 (C7) locus. To determine if this translocation caused the Rett s yndrome in this patient, our efforts focused on mapping and cloning of the X chromosomal breakpoint in this patient. Toward these goals, we generated a set of radiation-reduced hybrid cell lines for the short a rm of the X chromosome to use as a source for region-specific markers. Using Alu-PCR, 13 new DNA markers were isolated from a radiation-redu ced hybrid, which retained both DMD and DXS28. These markers were loca lized within Xp21 using DNA from males with various interstitial delet ions in this region. Two new markers, K23-2p and K23b-1, were found to be closer flanking markers to the X chromosomal breakpoint than DMD a nd DXS28. Long range restriction mapping using K23-2p and K23b-1 deter mined that the maximum distance between them was 800 kb. Several of th e new markers were developed into sequence tagged-sites and were used to isolate yeast artificial chromosome (YAC) clones. A total of 22 YAC clones was isolated and characterized; these YACs were then developed into 3 large contigs in the Xp21.3 region. This effort resulted in th e cloning of the region containing the X chromosomal translocation bre akpoint of the Rett syndrome patient in a 170.kb YAC clone. (C) 1993 W iley-Liss, Inc.