Targeted 'Next-Generation' Sequencing in Anophthalmia and Microphthalmia Patients Confirms SOX2, OTX2 and FOXE3 Mutations
Lopez Jimenez, Nelson
Sherr, Elliott H
Slavotinek, Anne M
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CitationLopez Jimenez, Nelson, Jason Flannick, Mani Yahyavi, Jiang Li, Tanya Bardakjian, Leath Tonkin, Adele Schneider, Elliott H Sherr, and Anne M Slavotinek. 2011. Targeted 'Next-Generation' Sequencing in Anophthalmia and Microphthalmia Patients Confirms SOX2, OTX2 and FOXE3 Mutations. BMC Medical Genetics 12:172.
AbstractBackground: Anophthalmia/microphthalmia (A/M) is caused by mutations in several different transcription factors, but mutations in each causative gene are relatively rare, emphasizing the need for a testing approach that screens multiple genes simultaneously. We used next-generation sequencing to screen 15 A/M patients for mutations in 9 pathogenic genes to evaluate this technology for screening in A/M. Methods We used a pooled sequencing design, together with custom single nucleotide polymorphism (SNP) calling software. We verified predicted sequence alterations using Sanger sequencing. Results: We verified three mutations - c.542delC in SOX2, resulting in p.Pro181Argfs*22, p.Glu105X in OTX2 and p.Cys240X in FOXE3. We found several novel sequence alterations and SNPs that were likely to be non-pathogenic - p.Glu42Lys in CRYBA4, p.Val201Met in FOXE3 and p.Asp291Asn in VSX2. Our analysis methodology gave one false positive result comprising a mutation in PAX6 (c.1268A > T, predicting p.X423LeuextX*15) that was not verified by Sanger sequencing. We also failed to detect one 20 base pair (bp) deletion and one 3 bp duplication in SOX2. Conclusions: Our results demonstrated the power of next-generation sequencing with pooled sample groups for the rapid screening of candidate genes for A/M as we were correctly able to identify disease-causing mutations. However, next-generation sequencing was less useful for small, intragenic deletions and duplications. We did not find mutations in 10/15 patients and conclude that there is a need for further gene discovery in A/M.
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