Exome Sequencing from Blood DNA

DNA from blood was submitted to The Center for Applied Genomics (TCAG), Toronto, Canada for exome library preparation and sequencing. DNA was quantified by Qubit DNA HS assay (Life Technologies, Carlsbad, CA) and 100 ng of input DNA was used for library preparation using the Ion AmpliSeq Exome Kit (Life Technologies) according to the manufacturer's recommendations. The Ampliseq Exome library was immobilized on Ion PI™ Ion Sphere™ particles using the Ion PI Template OT2 200 Kit v3. Sequencing was performed with the Ion PI Sequencing 200 Kit v3 and Ion PI Chip v2 in the Ion Proton™ semiconductor sequencing system following the manufacturer's recommendation. Alignment and variant calling were performed using Torrent Suite (v4.0) on the Ion Proton Server, using the Ion Proton AmpliSeq germline low stringency setting and the hg19 reference genome. The variants were annotated using an in-house annotation pipeline (22 (link)) based on Annovar (November 2014 version) (23 (link)) and RefSeq gene models (downloaded from UCSC 01 August 2015).