A complete workflow from sample preparation to analysis using SureSelect target enrichment system for Ion Proton semiconductor sequencing

Whole exome or targeted sequencing for protein-coding regions has provided a cost effective way to identify common and rare polymorphisms that are associated with Mendelian disorders and complex diseases. With increased capacity of semiconductor sequencing, highly multiplexed samples can be studied in a single sequencing run. However, a complete workflow processing raw DNA samples to identify DNA variants in target regions is not easily accessible. Here we describe an analysis workflow to study multiplexed samples in semiconductor sequencing for several target sizes: 50Mb (Human All Exon), 3.2Mb (Human Kinome) and a 1Mb custom design. The workflow includes library preparation, SureSelect target enrichment, semiconductor sequencing, and variant calling with SureCall software (beta version). Improved and simplified steps for library preparation and target enrichment maximize multiplexing and produce consistent results in the Ion Proton sequencer. Sequencing output can be easily analyzed, visualized and summarized in a report with SureCall which is optimized for use with Agilent’s target enrichment system. We demonstrate high capture efficiency, uniformity, and reproducibility of enrichment. The results from different capture sizes show comparable high performance regardless of various targeted regions. The combination of efficient target enrichment system, semiconductor sequencing, and SureCall software provides a fast and convenient tool to assess DNA variants in genomic regions of
interest.