Semiconductor Sequencing of Human Exomes on the Ion Proton System

Rapid, accurate, and inexpensive sequencing of exomes is critical to understand DNA variation in human disease. Ion Torrent has developed a benchtop research semiconductor sequencer, the Ion Proton™, that uses a novel CMOS chip with 165 million 1.3mm-diameter microwells, automatically templated sub-micron particles, and integrated hardware and software that enables acquisition of ~5 billion data points per second over a 2-4 hour runtime with on-instrument signal processing. To illustrate the speed, accuracy, and ease-of-use of the Proton system, analysis of a HapMap familial trio of exomes will be presented. Exome libraries are obtained with high-specificity hybridization probes targeting ~50 Mb of human exons that span 21,700 annotated protein-coding genes, microRNA, key non-coding RNA genes, and 44,000 predicted microRNA binding sites. Exome reads map on-target 75-83% between runs and 10.6 Gb of aligned data, obtained from a single P1 chip, yielded 141X average depth with 30X coverage of 90% of targeted bases. Read mapping, coverage analysis, variant calling and annotation are done with Torrent Suite and Ion Reporter™ software. Each trio dataset yielded ~30,000 SNP calls from single runs that exceeded 9 Gb of aligned data. The observed Het:Hom ratio of 1.4-1.5 matches the published range of 1.25-1.7 for European ethnicity and the observed Ts:Tv ratio of 2.9 agrees well with the published range of 2.8-3.1 for human exomes. The SNP concordance with dbSNP137 is greater than 98% and Het and Hom concordances with Complete Genomics data are 98% and 96%, respectively. Mendelian inheritance analysis indicates that error for Hets is 0.6% with no errors for homozygotic SNPs. The Proton system delivers high-quality individual exome datasets rapidly and can be used for trio analysis to detect shared germline SNPs with high confidence. The Ion Proton™ System is for research use only and not for use in diagnostic procedures.