外显子组测序PPT.pptx

外显子组测序目 录一、外显子测序简介 外显子测序（也称目标外显子组捕获）是指利用序列捕获技术将全基因组外显子区域DNA捕捉并富集后进行高通量测序的基因组分析方法。是一种选择基因组的编码序列的高效策略，外显子测序相对于基因组重测序成本较低，对研究已知基因的SNP、Indel等具有较大的优势。 在人类基因中大约有180,000外显子，占人类基因组的1%，约30MB。Ng S B, Turner E H, Robertson P D, et al. Targeted capture and massively parallel sequencing of 12 human exomes[J]. Nature, 2009, 461(7261): 272-276. 人类基因组的蛋白编码区域大约包含85%的致病突变。- Choi M, Scholl U I, Ji W, et al. Genetic diagnosis by whole exome capture and massively parallel DNA sequencing[J]. Proceedings of the National Academy of Sciences, 2009, 106(45): 19096-19101.二、测序深度The sensitivity to detect heterozygous variants with 10 reads is 78.6%, but increases to 95.2% at 20x and approximately 100% at 30x and greater.[1]The average cover-age of each base in the targeted regions was 100-fold, and 95.3% of these bases were covered sufficiently deeply for variant calling (≥10× cover-age) [2]Exome sequencing produced a higher level of coverage for the targeted sequences (mean, 167.50×), slightly increasing our ability to detect mutations with VAFs of less than 10%. [3]Choi M, Scholl U I, Ji W, et al. Genetic diagnosis by whole exome capture and massively parallel DNA sequencing[J]. Proceedings of the National Academy of Sciences, 2009, 106(45): 19096-19101.Yan X J, Xu J, Gu Z H, et al. Exome sequencing identifies somatic mutations of DNA methyltransferase gene DNMT3A in acute monocytic leukemia[J]. Nature genetics, 2011, 43(4): 309-315.Platforms A. Genomic and Epigenomic Landscapes of Adult De Novo Acute Myeloid Leukemia[J]. N Engl J Med, 2013, 2013(368): 2059-2074.基于Ion Proton?的外显子测序流程The bound DNA is isolated using streptavidin-coated Dynabeads? paramagnetic beads, and then amplified and purified. The purified, target-enriched sample is then returned to the Ion Torrent system workflow for emulsion PCR, enrichment, and sequencing.Exome sequencing results on the Ion Proton? System using the Ion PI? Chip and the Ion TargetSeq? Exome Kit基于Ion Proton?的外显子测序结果Raw reads Reads mapped Percent reads m