Nonsynonymous variants in MYH9 and ABCA4 are the most frequent risk loci associated with nonsyndromic orofacial cleft in Taiwanese population

BACKGROUND: Nonsyndromic orofacial cleft is a common birth defect with a complex etiology, including multiple genetic and environmental risk factors. Recent whole genome analyses suggested associations between nonsyndromic orofacial cleft and up to 18 genetic risk loci (ABCA4, BMP4, CRISPLD2, GSTT1, FGF8, FGFR2, FOXE1, IRF6, MAFB, MSX1, MTHFR, MYH9, PDGFC, PVRL1, SUMO1, TGFA, TGFB3, and VAX1), each of which confers a different relative risk in different populations. We evaluate the nonsynonymous variants in these 18 genetic risk loci in nonsyndromic orofacial clefts and normal controls to clarify the specific variants in Taiwanese population. METHODS: We evaluated these 18 genetic risk loci in 103 cases of nonsyndromic orofacial clefts and 100 normal controls using a next-generation sequencing (NGS) customized panel and manipulated a whole-exon targeted-sequencing study based on the NGS system of an Ion Torrent Personal Genome Machine (IT-PGM). IT-PGM data processing, including alignment with the human genome build 19 reference genome (hg19), base calling, trimming of barcoded adapter sequences, and filtering of poor signal reads, was performed using the IT platform-specific pipeline software Torrent Suite, version 4.2, with the plug-in “variant caller” program. Further advanced annotation was facilitated by uploading the exported VCF file from Variant Caller to the commercial software package Ion Reporter; the free online annotation software Vanno and Mutation Taster. Benign or tolerated amino acid changes were excluded after analysis using sorting intolerant from tolerant and polymorphism phenotyping. Sanger sequencing was used to validate the significant variants identified by NGS. Furthermore, each variant was confirmed in asymptomatic controls using the Sequenom MassARRAY (San Diego, CA, USA). RESULTS: We identified totally 22 types of nonsynonymous variants specific in nonsyndromic orofacial clefts, including 19 single nucleotide variants, 2 deletions, and 1 duplication in 10 studied genes(ABCA4, MYH9, MTHFR, CRISPLD2, FGF8, PVRL1, FOXE1, VAX1, FGFR2, and IRF6). Nonsynonymous variants in MYH9 and ABCA4, which were detected in 6 and 5 individuals, respectively, were identified to be the most frequent risk loci in nonsyndromic orofacial clefts in the Taiwanese population. CONCLUSIONS: Nonsynonymous variants in MYH9 and ABCA4 were identified to be the most frequent risk loci in nonsyndromic orofacial clefts in the Taiwanese population. These findings in our study have provided additional information regarding specific variants associated with nonsyndromic orofacial clefts in different population and demonstrate the power of our customized NGS panel, which is clinically useful for the simultaneous detection of multiple genes associated with nonsyndromic orofacial clefts. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1186/s12881-016-0322-2) contains supplementary material, which is available to authorized users.