An integrated physical, genetic and transcript map will be constructed for the Xq21.1-q21.3 region on the long arm of the human X chromosome. This region encompasses approximately 15-20 megabases (Mb) of the physical length of the X chromosome. The physical map will be constructed with Yeast artificial chromosome (YAC) contigs isolated using over 30 DNA . probes and genes located in the Xq21.1-q21.3 region and 33 STSs defining the X/Y homology region. The ICRF and other human YAC libraries will be screened by both hybridisation and PCR and bidirectional chromosome walks will be used to fill in the gaps. YACS will be tested by FISH on normal chromosomes for chimeric inserts and on a subset of cell-lines derived from female patients with X;autosome translocations and males with deletions breaking in this region to define the physical relationship of YACs to the breakpoints. All genes, DNA Probes, STSs, YAC end-probes, and new microsatellite markers generated in this project will be tested by PCR analysis or hybridisation on a Series of 9 translocation hybrid cell-lines derived from females with X;autosome translocations, and 17 lymphoblastoid cell-lines from males with deletions and duplications that break the Xq21 region into 19 intervals. To develop new genetic markers in this region, YACs will be either hybridised to the ICRF human X chromosome cosmid grids or subcloned into cosmid and smaller insert sublibraries and short tandem repeat polymorphisms (microsatellites) will be isolated and tested in standard CEPH pedigrees. The transcript map of the region will be constructed using cosmid contigs generated from the YAC clones in each contig. Genes will be identieied mostly using 1) Exon amplification of cosmids and 2) CDNA selection procedures using hybridisation of PCR amplified CDNA library inserts to pooled biotinylated cosmid DNA. Transcribed sequences isolated from the region will be further analyzed for tissue distribution of expression and related to several disease loci mapping in deletion intervals in this region (X-linked deafness, mental retardation, and cleft lip and palate).
Funding SchemeCSC - Cost-sharing contracts