Integrated Bioinformatics Analysis Suite for deep, targeted amplicon sequencing

描述（由申请人提供）：本提案的目的是开发一个生物信息学管道，或集成生物信息学分析套件（），专门用于在新的Ion Torrent个人基因组机（PGM）上进行超深度靶向扩增子测序。这将提高变异检测的准确性，并使分析不同类型癌症的遗传变异成为可能。PGM中使用的技术是基于半导体技术的非光学检测，因此，当其他技术用于初始测序时，它足以作为验证性测试的正交平台。通过使样品制备到数据生成的过程在一天内完成，PGM提供了比其他下一代测序方法显著的优势。然而，它受到技术固有的限制，主要是在32端错误。此外，目前可用的SNP呼叫者并不特别适合高通量，超

DESCRIPTION (provided by applicant): The aim of this proposal is to develop a bioinformatics pipeline, or Integrated Bioinformatics Analysis Suite (IBAS), specific for ultra-deep, targeted amplicon sequencing on the new Ion Torrent Personal Genome Machine (PGM). This will improve the accuracy of variation detection and enable analysis of genetic variations in different types of cancer. The technology used in the PGM is based on non-optical detection with semiconductor technology and so is sufficiently unique to enable its use as an orthogonal platform for confirmatory testing when other technologies are used for initial sequencing. The PGM offers a significant advantage over other methods of next generation sequencing by enabling the process of sample preparation to data generation to be completed in one day. However, it suffers from the inherent limitations of the technology, mainly errors at the 32 end. Additionally, currently available SNP callers are not particularly suited to high-throughput, ultra deep amplicon sequencing and there are few algorithms available for detecting other types of genomic variations. The proposed pipeline aims to address these issues and fill the urgent need for bioinformatics optimized for the panel-designed amplicon sequencing technology employed in the PGM. To achieve the proposed aims, 1) an IBAS that incorporates different approaches to read preprocessing, alignment to reference sequences, and error modeling to enable detection of single nucleotide polymorphisms (SNPS), insertion/deletions (indels), copy number variation (CNV) and other structural variation will be constructed; and 2) a comprehensive evaluation of the sources of variation will be performed, and the IBAS optimized for amplicon sequencing. The optimized pipeline will then be validated by processing data from the PGM for orthogonal validation of mutations detected in clinical thyroid fine needle aspiration biopsies. Successful outcome will enable Asuragen to provide a high-throughput ultra-deep sequencing service for clinical research and diagnostics. PUBLIC HEALTH RELEVANCE: We aim to develop a bioinformatics application specific for the technology employed in the recently launched Ion Torrent Personal Genome Machine (PGM) that will improve the accuracy of detecting genetic variations in different types of cancer. The PGM is a compact, bench-top next generation sequencing instrument that enables rapid sequencing of an entire genome and at a much lower cost than established systems. The application will be used by Asuragen to offer a high-throughput sequencing service for clinical research and diagnostics.