Keen, Thomas Jeffrey; (1996) Molecular genetic analysis of human inherited retinal degeneration. Doctoral thesis (Ph.D), UCL (University College London).

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Abstract

The aim of the work described in this thesis was to investigate human inherited retinal degenerations using the techniques of molecular genetics. The largest part of this, was a study of a particular type of autosomal dominant retinitis pigmentosa (adRP) with the ultimate objective of identification of the defective gene. This condition is characterised by a progressive loss of peripheral vision often leading to complete blindness. Other forms of adRP are known to result from mutations in two known genes and at a further six as yet uncharacterised loci. The form studied here was that which had previously been mapped to chromosome 7p. The first stage of this analysis was the refinement of the genetic location to an interval between the markers D7S484 and D7S795. After the disease locus was placed in this discrete interval, work commenced on a physical clone contig of the region. This was achieved using overlapping YAC clones, which were identified by STS content using microsatellite markers. STSs generated from the ends of the YAC clones were used to further refine the order of elements within the contig. The resulting contig spanned approximately 4.8 Mb of DNA including 2.8 Mb across the RP interval. Some features of the contig are discussed including a regional duplication and the initial stages of a transcriptional map. In addition, a mutation screen was made of the gene coding for the photoreceptor specific protein peripherin-RDS, this gene located on chromosome 6p is known to be involved in a range of retinal degeneration phenotypes. The coding region of the gene was screened in a total of 23 families affected by various forms of macular dystrophy. A single nonsense mutation was identified in one family, the mutation comprising the duplication of a 4 base pair element which was predicted to result in the creation of a termination codon and a potentially truncated protein. Finally the possible involvement of expanded (CAG)n repeats in adRP has been investigated using the technique of "repeat expansion detection". These were studied because of their known involvement in some neurodegenerative disorders, the retina being neural in origin seemed another possible target tissue for the effects of expansions to be apparent. Nine different families were studied, five of which were of types whose disease mapped to known chromosomal locations. No expansions were detected that co-segregated with disease, though in one family an expansion was observed in several generations that was apparently unconnected with the phenotype. Expansions of (CAG)n therefore be ruled out as a direct cause of the RP in these families. However it remains possible that repeat expansions are involved in some forms of retinal degeneration.

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