Variant annotations, filtering and prioritizing
The bioinformatics analysis began with the sequencing data. First, joint contamination and low quality variants (read depth < 15 and genotype quality score < 20) were removed. The reads were mapped to the reference human genome (UCSC GRCh37/hg19) using BWA (Burrows-Wheeler Aligner) software. After that, variants calls were conducted by the GATK (v3.7). Finally, ANNOVAR tool was applied to perform a series of annotations, such as the frequency in the databases, conservative prediction and pathogenicity prediction with available website tools for variants.
The work flow of quality control were carried out as shown in Figure 1. First, we remove variants with a MAF more than 0.01 in the 1000 Genomes Project (http://www.internationalgenome.org/), Exome Aggregation Consortium (ExAC) (http://exac.broadinstitute.org/), dbSNP (https://www.ncbi.nlm.nih.gov/snp/) in databases. Second, missense, damaging or loss of function variants of ABC genes were embraced for the subsequent analyses. Third, we concentrated on novel variants which were filtered by NCBI and Ensembl. In addition, we prioritize to pay attention to the novel variants that would likely have functional effect. For instance, a variant will be highlighted when it predicted to be simultaneously deleterious by SIFT, Mutation Taster and FATHMM software.