Journal
JOURNAL OF MEDICAL GENETICS
Volume 48, Issue 3, Pages 145-151Publisher
BMJ PUBLISHING GROUP
DOI: 10.1136/jmg.2010.083568
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Funding
- Health and Social Care Northern Ireland R&D of the Public Health Agency [RRG 4.43]
- Public Health Agency [RRG/3237/05] Funding Source: researchfish
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Background The genetic heterogeneity of many Mendelian disorders, such as retinitis pigmentosa which results from mutations in over 40 genes, is a major obstacle to obtaining a molecular diagnosis in clinical practice. Targeted high-throughput DNA sequencing offers a potential solution and was used to develop a molecular diagnostic screen for patients with retinitis pigmentosa. Methods A custom sequence capture array was designed to target the coding regions of all known retinitis pigmentosa genes and used to enrich these sequences from DNA samples of five patients. Enriched DNA was subjected to high-throughput sequencing singly or in pools, and sequence variants were identified by alignment of up to 10 million reads per sample to the normal reference sequence. Potential pathogenicity was assessed by functional predictions and frequency in controls. Results and conclusions Known homozygous PDE6B and compound heterozygous CRB1 mutations were detected in two patients. A novel homozygous missense mutation (c.2957A -> T; p.N986I) in the cyclic nucleotide gated channel beta 1 (CNGB1) gene predicted to have a deleterious effect and absent in 720 control chromosomes was detected in one case in which conventional genetic screening had failed to detect mutations. The detection of known and novel retinitis pigmentosa mutations in this study establishes high-throughput DNA sequencing with DNA pooling as an effective diagnostic tool for heterogeneous genetic diseases.
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