Targeted Sequencing of ST18 Gene

DNA enrichment was performed using HaloPlex kit (Agilent Technologies, Santa Clara, CA, USA) and sequencing was conducted on a MiSeq system sequencer (Illumina, San Diego, CA, USA) with 150 bp paired-end reads. A total of 463407 bp were included in the capture design, covering the entire ST18 gene (chr8: 53,023,399–53,373,519, GRCh37/hg19 assembly) as well as 10 kb downstream and 50 kb upstream to the gene and an additional 2 Mb located upstream and downstream to the gene and predicted to harbor putative regulatory regions (https://www.encodeproject.org).
The sequencing data were processed using MiSeq Reporter 2.0.26 and Casava softwares (Illumina, San Diego, CA, USA) and analyzed for quality control using FastQC software (http://www.bioinformatics.babraham.ac.uk/projects/fastqc). Reads were aligned to the Genome Reference Consortium Human Build 37 (GRCh37/hg19) using Burrows-Wheeler Aligner [40 (link)] and variant detection was achieved using The Genome Analysis Toolkit [41 (link)]. Variants were annotated by ANNOVAR [42 (link)] and the frequency of each variant was determined using data from dbSNP138, the 1000 Genome Project and an in-house database. Case-control association test for variants was performed with chi-square, and permutation test, using the Caucasian population from the 1000 Genome Project data (http://www.1000genomes.org) as a control.