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Derek Le, Doug Chung, Ricardo F Frausto, Michelle Jangmi Kim, Anthony J Aldave; Identification of potentially pathogenic noncoding region variants in the posterior polymorphous corneal dystrophy 1 locus. Invest. Ophthalmol. Vis. Sci. 2015;56(7 ):2521.
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© ARVO (1962-2015); The Authors (2016-present)
To identify the genetic basis of posterior polymorphous corneal dystrophy 1 (PPCD1) using a next-generation sequencing (NGS) approach.
NGS was performed on DNA samples from 4 affected and 4 unaffected members of a family previously linked to a 14 Mb region on chromosome 20 between markers D20S182 and D20S195, encompassing the PPCD1 locus. Custom probes were used for targeted region capture of the linked interval. The exons, splice sites, 5’ and 3’ UTRs, and promoter regions (defined as -1 Kb from transcription start site) of all annotated protein-coding genes within the captured region were examined for sequence variants. Single nucleotide variants (SNVs) and insertions/deletions (indels) were called using Bowtie 2 for alignment and SAMtools for variant identification. An independent indel analysis was performed using Burrows Wheeler Aligner for alignment and HaplotypeCaller (HC) for variant identification. Quality (≥20) and read depth (≥5X) filters were applied to generate lists of all variants. Potentially pathogenic (candidate) variants met the following criteria: heterozygous, nonsynonymous, novel or rare (minor allele frequency (MAF) < 0.05), and present in each of the affected and none of the unaffected individuals.
An average of ~12,900 SNVs were identified per individual. No coding region candidate SNVs were identified. One noncoding candidate SNV was identified: a novel variant in the promoter region of OVOL2 (c.-307T>C). An average of ~1,200 and ~1,400 indels were identified per individual with SAMtools and HC, respectively. No coding region candidate indels were identified with either SAMtools or HC. Two noncoding candidate indels were identified: a novel insertion in the 5’ UTR of RIN2 (c.-55_-54insCTT) using SAMtools and a known (but with unknown MAF) insertion in the promoter region of OVOL2 (c.-769insC) using HC.
Using a NGS approach to identify SNVs and indels in the PPCD1 locus, no potentially pathogenic coding region variants were identified. However, potentially pathogenic variants (one SNV and two indels) were identified in the noncoding regions of two genes, OVOL2 and RIN2. Screening for these variants in other affected and unaffected family members, as well as in the noncoding regions of these genes in other PPCD1 probands, will determine which, if any, is associated with PPCD1.
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