The development and application of SNP discovery technologies from 2005 to 2012

Colley, James P. M. 2012. The development and application of SNP discovery technologies from 2005 to 2012. PhD Thesis, Cardiff University. Item availability restricted.

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Abstract

Single Nucleotide Polymorphisms (SNPs) are used as markers in association studies and may contribute directly to inherited disease. Here, we investigated high throughput in silico and in vitro methods for identifying SNPs and applied these to large scale genomic projects. We evaluated the utility of the Transgenomic NavigatorTM software to facilitate automated detection of aberrant denaturing high performance liquid chromatography (dHPLC) elution profiles. 3,747 dHPLC profiles were analysed with this software and 98.3% of products with profiles distinct from wild type harboured novel variants (Chapter 3). We applied this software to investigate whether rare inherited variants in genes that play a role in oxidative DNA damage repair (OxDR) predispose to colorectal adenomas (CRA) and found that unlike MUTYH, inherited variants in other OxDR genes are unlikely to be a frequent cause of CRA (Chapter 4). We evaluated the sequence analysis packages Sequencher, InSNP, Mutation Surveyor and Staden to identify variants in patients with CRAs (using >4,000 chromatograms). Staden and Mutation Surveyor correctly identified 76/77 (98.7%) SNPs and 96.7% and 99.3% of the genotypes respectively (Chapter 5). We compared an optimised version of Staden against Sequencher for variant detection over a 2.5kb region of the adenomatous polyposis coli (APC) gene in 969 healthy controls. We found 100% concordance between these packages and found that rare nonsynonymous variants in APC were significantly over-represented in patients with CRAs as compared to healthy controls (32/480 vs. 37/969, P=0.0166) (Chapter 5). We evaluated data held in dbSNP (build 129) for common variants in the Caucasian population by examining ten DNA repair genes and subsequently developed software for automatically extracting selected information from dbSNP (Chapter 6). This program was used to rapidly identify 221 common nonsynonymous SNPs in every DNA repair gene in the human genome; these were subsequently typed in 480 publically-available human lymphoblastoid cell lines generating a resource for functional analyses (Chapter 6). We also assessed the role of these SNPs in susceptibility to CRC and response to therapy by exploiting data and samples for the randomised controlled trial COIN (Chapter 7). Finally, we used Next Generation Sequencing (NGS) to discover SNPs in the exomes of 10 patients that exhibited peripheral neuropathy in response to chemotherapy and discovered that ERCC4 nonsense mutations may contribute to this toxicity (Chapter 8). NGS is likely to become the key SNP discovery technique over the next decade due to its potential to comprehensively search an entire genome at a comparatively low cost.

Item Type:	Thesis (PhD)
Status:	Unpublished
Schools:	Medicine
Subjects:	R Medicine > R Medicine (General)
Date of First Compliant Deposit:	30 March 2016
Last Modified:	12 Jun 2019 02:16
URI:	https://orca.cardiff.ac.uk/id/eprint/41990

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