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Mansoor Sarfarazi, Roshanak Sharafieh, Robert Ritch, Jeffrey M Liebmann, Ahti Tarkkanen, Francesc Lopez-Giraldez, Kaya Bilguvar, Richard P Lifton, Shrikant Mane, Anne H Child; Exome Sequencing of Familial and Sporadic Cases with Pseudoexfoliation Syndrome (XFS). Invest. Ophthalmol. Vis. Sci. 2014;55(13):3815. doi: https://doi.org/.
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© ARVO (1962-2015); The Authors (2016-present)
To identify new causative genes in a group of XFS subjects by next-generation exome sequencing (NGES) method.
The cohort consisted of 72 affected XFS individuals, including 28 members of 13 unrelated families (Sibs, Parent-Child, Niece-Uncle, 1st to 3rd degree cousins) and 44 sporadic cases (28 with and 16 without Glaucoma). Exome capture was performed using the NimbleGen SeqCap EZ V2 followed by sequencing on the Illumina HiSeq 2000. Reads were mapped to the human genome (hg19) with ELAND aligner and variants were called with SAMtools.
Exome sequencing revealed 2,539,812 variants in 72 XFS subjects, of which 90% had a Quality Score of ≥60 and 94% of bases were covered at least 8-times. Removal of duplicates and common alterations revealed 470,441 variants in 17,989 unique genes. Subsequent filtrations against 1,000 Genome, NHLBI Exomes, dbSNP, Simple Repeats and Genome Segmental Duplications identified 291,022 novel variants, of which 37,501 (8,347 Missense; 1,631 Nonsense; 7,022 Synonymous; 20,501 Non-Coding) were present in 2 or more affected XFS subjects. Overall, 113 genes showed novel amino acid alterations in at least 10% of our XFS cases. However, when familial co-segregation of these novel coding variants was considered, only 12 genes segregated in one or more of our XFS families. Bioinformatic analysis and direct-Sanger sequencing are currently being utilized to evaluate the causative nature of these candidate genes in our original exome-sequenced subjects and their immediate respective family members. Additionally, we have observed another 24,281 insertions or deletions (InDels) in our 72 XFS subjects, of which 52 novel variants were identified in 2 or more affected cases. However, only 22 of these InDels were observed in both familial and sporadic cases. The exact contribution of these InDels and their roles in the etiology of XFS phenotype is currently being evaluated.
This is the first study to use NGES in a group of 72 familial and sporadic XFS cases. The NGES identified 12 genes with single novel amino acid alterations and 22 other genes with InDels predicting to instigate premature termination of their encoded normal proteins. The exact nature of these DNA alterations and their potential XFS-causing roles are currently under investigation. Note: Other contributing authors are M.E. Turacli, C. O’Brien, F. Mercieca and A. Spiteri
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