Cytosnp850k array

For the paired-end pre-capture library procedure, genomic DNA from a lesional skin and blood was fragmented by sonication and ligated to the Illumina multiplexing paired-end (PE) adapters. The adapter-ligated DNA was then polymerase chain reaction (PCR) amplified using primers with sequencing barcodes (indexes). For target enrichment/exome capture procedure, the pre-capture library was enriched by hybridizing to biotin labeled VCRome 2.1 in-solution exome probes (Bainbridge et al., 2011 (link)) for 64–72 h at 47°C. Additional probes for over 3600 Mendelian disease genes were also included in the capture in order to improve the exome coverage. For massively parallel sequencing, the post-capture library DNA was subjected to sequence analysis on Illumina HiSeq platform for 100-bp paired-end reads. The following quality control metrics of the sequencing data are generally achieved: >70% of reads aligned to target, >95% target base covered at >20X, >85% target base covered at >40X, mean coverage of target bases >100X. Single-nucleotide polymorphism (SNP) concordance to genotype array: >99%. As a quality control measure, DNA was also analyzed by a SNP array: Illumina Human Exome-12v1 array and the Illumina CytoSNP-850K array. The SNP data are compared with the WES data to ensure correct sample identification and to assess sequencing quality.