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Terry Gaasterland, Douglas E Gaasterland, Lee E Edsall, Amy N Dubinsky, Kuan-Fu Ding, Steven Head, Karl H Willert, Consortium The NEIGHBOR, ; Identification of disease-associated genome variants in regulatory regions using exome sequencing in 295 POAG cases. Invest. Ophthalmol. Vis. Sci. 2014;55(13):3792.
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© ARVO (1962-2015); The Authors (2016-present)
To discover disease-associated genome variants in coding or regulatory regions of genes.
DNA collected from 295 affected individuals was enriched using probes designed to capture coding exons, untranslated regions (UTR), flanking intron, and proximal promoters. Patient samples were a subset from the NEIGHBOR GWAS, all with high pressure primary open angle glaucoma (POAG). DNA was sequenced on Illumina instruments, generating at least 10 gigabases of data per sample. Reads were mapped to the hg19 reference human genome and single nucleotide polymorphisms (SNPs) identified using methods published in the literature with parameters optimized to minimize false positives. Allele and zygosity frequencies were calculated from genotype calls. Comparisons between cases and available general population databases were based on the less frequent (minor) allele in 1000G for every SNP site identified in at least 30 patients. For annotation, lists were constructed with genes reported as involved in glaucoma, retinal disease and other neurodegenerative diseases or with enriched expression in optic tissue.
Clinical considerations allowed definition of constraints to apply to the SNP sites including the following: (a) A genomic aberration is not likely to be important as a primary cause if it occurs with frequency close to that in a general population; so a site was excluded unless its frequency in the cases was at least 0.20 higher than in every comparison database. (b) If a high frequency anomaly were causing disease, it would be more prevalent in the general population; so, although there is a possibility of lack of penetrance, sites with allele frequency >0.25 in any general population were excluded. Fisher's exact test with correction for multiple testing was used to assign p-values to resulting sites. This yielded 165 variants in 99 genes with significant p-values.
This search used SNPs found in codon, UTR, intron, and promoter regions, and revealed potentially causative genes related to this genetically complex, chronic eye disease. Genome variations with markedly higher occurrence in high pressure POAG patients than in the general population implicated 99 genes of the approximately 25,000 genes encoded in the human genome. This study supplements recent advances in understanding of POAG based on gene implication from GWAS and GWAS-related pathway analyses.
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