Genome Studies of Human Chromosome 22q13.3.

This thesis describes my work in constructing a Y AC framework map and subsequent bacterial clone contig of human chromosome 22ql3.3. The region was spanned by 5 conrigs containing 1986 bacterial clones, 84 minimal overlapping clones were sequenced. The integration of landmark content data allowed the ordering of contigs. The 4.3 Mb sequence map was used to assemble a transcript map across 22ql3.3. The sequence allowed the annotation of 47 genes and 6 pseudogenes. In addition, the genes’ environment was studied. Assessment of the repeat content showed that nearly 30 percent was repetitive. A strong inverse correlation was observed between Line 1 density and GC content. No correlation was observed for Alu density and GC content. The gene density was shown to increase with higher GC content. Assessment of splice sites showed strong conservation of splice donor and acceptor sites. The protein products were characterised by an integrated computational approach, the results were graphically viewable using the peptide display of ACeDB. For a 140 kb region, containing 9 genes, at the telomeric end of 22ql3.3 a mouse comparative map was constructed and a single clone sequenced. The comparison of the mouse and human sequence showed complete conservation of gene order and content. Strong conservation was observed of splice sites and protein coding regions. The utility of comparative sequencing was shown by identification of a previously missed gene. Redundant sequencing of this same region also allowed the identification of 220 candidate variations. Of these 12 were verified by PCR-RFLP and all were shown to be polymorphic. Extrapolation of this data suggests that most of the identified variations are likely to be polymorphic. As expected a strong bias was observed towards transversions, but the data also showed a negative bias with regard to A to T or T to A changes