Targeted Sequencing of Hearing Loss Genes

Genomic DNA (gDNA) was extracted from the whole blood samples using the Blood DNA kit (TIANGEN BIOTECH, Beijing, China), and 1 ug of purified gDNA fragmented to 200–300 base pairs using an ultrasonoscope (Covaris S2, Massachusetts, USA). End-repair, adenylation and adapter ligation were performed for library preparation following the Illumina's protocol. The same amount of library were pooled then hybridized to the customized capture array (NimbleGen, Roche) including exons, splicing sites and immediate flanking intron sequences of 29 genes for non-syndromic autosomal dominant hearing loss and TNC, a novel causative gene for ADNSHL identified in our previous research (Table S1). Sequencing was carried out on Illumina HiSeq2000 to generate paired end reads (90 bps at each end) [7] (link).
Raw image files were processed by Illumina Pipeline (version 1.3.4) for base-calling with default parameters. Reads were aligned to NCBI37/hg19 assembly using the BWA (Burrows Wheeler Aligner). SNPs and indels (inserts and deletions) were detected using the GATK software [8] (link).