ISDT:一种基于高通量测序数据的外源序列插入位点检测工具  

ISDT:a Tool for Detecting Exogenous Sequence Insertion Sites based on Highthroughput Sequencing Data

作　　者：张镇 杨芊茹 杜含笑 李雨桐 陆晨琪 姜宁 ZHANG Zhen;YANG Qianru;DU Hanxiao;LI Yutong;LU Chenqi;JIANG Ning(School of Life Sciences,Fudan University,Shanghai,200433;WuXi Biologics Biosafety Testing(Suzhou)Co.,Ltd.,Suzhou,215000)

机构地区：[1]复旦大学生命科学学院,上海200433 [2]苏州药明检测检验有限责任公司,苏州215000

出　　处：《基因组学与应用生物学》2024年第9期1510-1520,共11页Genomics and Applied Biology

基　　金：国家重点研发计划(2021YFC2501800,2022YFC2009802)资助。

摘　　要：精准识别外源序列插入位点对于转基因研究、基因编辑研究及疾病机制探索等领域至关重要。随着高通量测序技术的快速发展和广泛应用,越来越多的研究者将高通量测序技术应用到插入位点分析研究中来,深入探索外源序列插入事件。基于这样的背景,本文开发一款基于高通量测序数据的外源序列插入位点精准识别检测工具(insertion sites detection tool,ISDT),该工具引入了一种新颖的外源序列插入位点精准识别算法,该算法的核心是基于滑动打断候选read的比对和4种模式检索(Forward-Reverse模式、Reverse-Forward模式、Forward-Forward模式和Reverse-Reverse模式)从而实现精准识别插入位点。为评估该插入位点检测工具的性能,我们不仅采用不同插入类型(包括单点插入类型、单个外源片段插入类型、外源性同源片段插入类型、不同染色体上的外源性片段插入类型、不同染色体上的外源性同源片段插入类型)的模拟数据和植物T-DNA、动物F8基因的真实数据进行了全面的测试,还将该检测工具和发表的分析方法进行了比较,本工具均展现了出色的准确性和灵敏度。本文开发的检测工具作为一种高效、准确的插入位点检测工具,其跨物种的适用性及对真实复杂数据的处理能力,为未来的转基因研究、基因编辑研究及疾病机制探索等提供了重要的支持,有着广阔的应用前景。Accurate identification of exogenous sequence insertion sites is crucial for transgenic research,gene editing studies,and exploration of disease mechanisms.With the rapid development and widespread application of high-throughput sequencing technology,an increasing number of researchers are applying this technology to the analysis of insertion sites,delving into exogenous sequence insertion events.Based on this background,we developed a detection tool for precise identification of exogenous sequence insertion sites based on high-throughput sequencing data(insertion sites detection tool,ISDT).This tool introduces a novel algorithm for precise identification of insertion sites,which relies on alignment of sliding candidate reads and four mode searches(Forward-Reverse mode,Reverse-Forward mode,Forward-Forward mode,and Reverse-Reverse mode)to achieve accurate insertion site identification.To evaluate the analytical performance of this insertion site analysis tool,we not only used simulated high-throughput sequencing data containing different types of insertions(including single-point insertion,single exogenous fragment insertion,exogenous homologous fragment insertion,exogenous fragment insertion on different chromosomes,exogenous homologous fragment insertion on different chromosomes)and real insertion site high-throughput sequencing data of plant T-DNA and animal F8 gene for comprehensive testing,but also compared this analysis tool with published analysis methods.Our tool demonstrated excellent accuracy and sensitivity.The insertion site detection tool developed in this paper,as an efficient and accurate tool for precise identification of insertion sites,offers cross-species applicability and the ability to handle real complex data.This provides significant support for future transgenic research,gene editing studies,and the exploration of disease mechanisms,holding broad application prospects.

关 键 词：高通量测序 外源序列 插入位点 精准识别 检测工具 

分 类 号：Q811.4[生物学—生物工程]