Discovery of rare mutations in extensively pooled DNA samples using multiple target enrichment

doi:10.1111/pbi.12174

	Discovery of rare mutations in extensively pooled DNA samples using multiple target enrichment
	Chi, Xu 1; Zhang, Yingchun ; Xue, Zheyong ; Feng, Laibao ; Liu, Huaqing 2; Wang, Feng 2; Qi, Xiaoquan
	2014
发表期刊	PLANT BIOTECHNOLOGY JOURNAL
ISSN	1467-7644
卷号	12 期号:6 页码:709-717
摘要	Chemical mutagenesis is routinely used to create large numbers of rare mutations in plant and animal populations, which can be subsequently subjected to selection for beneficial traits and phenotypes that enable the characterization of gene functions. Several next-generation sequencing (NGS)-based target enrichment methods have been developed for the detection of mutations in target DNA regions. However, most of these methods aim to sequence a large number of target regions from a small number of individuals. Here, we demonstrate an effective and affordable strategy for the discovery of rare mutations in a large sodium azide-induced mutant rice population (F-2). The integration of multiplex, semi-nested PCR combined with NGS library construction allowed for the amplification of multiple target DNA fragments for sequencing. The 8 x 8 x 8 tridimensional DNA sample pooling strategy enabled us to obtain DNA sequences of 512 individuals while only sequencing 24 samples. A stepwise filtering procedure was then elaborated to eliminate most of the false positives expected to arise through sequencing error, and the application of a simple Student's t-test against position-prone error allowed for the discovery of 16 mutations from 36 enriched targeted DNA fragments of 1024 mutagenized rice plants, all without any false calls.
关键词	NGS induced mutation detection large samples pooling multiplexed target enrichment
学科领域	Biotechnology & Applied Microbiology ; Plant Sciences
DOI	10.1111/pbi.12174
收录类别	SCI
语种	英语
WOS关键词	POPULATIONS ; PCR ; POLYMORPHISMS ; AMPLIFICATION ; GENERATION ; GENES ; AZIDE
WOS研究方向	Biotechnology & Applied Microbiology ; Plant Sciences
WOS记录号	WOS:000339488200007
出版者	WILEY
文献子类	Article
出版地	HOBOKEN
EISSN	1467-7652
资助机构	National Transgenic Megaproject of China [2013ZX08009001] ; Chinese National Programs for Fundamental Research and Development [2013CB127000]
作者邮箱	xqi@ibcas.ac.cn
作品OA属性	Green Published, hybrid
引用统计	被引频次：9[WOS] [WOS记录] [WOS相关记录]
文献类型	期刊论文
条目标识符	http://ir.ibcas.ac.cn/handle/2S10CLM1/27271
专题	中科院植物分子生理学重点实验室
作者单位	1.Chinese Acad Sci, Inst Bot, Key Lab Plant Mol Physiol, Beijing, Peoples R China 2.Chinese Acad Sci, Grad Univ, Beijing, Peoples R China 3.Fujian Acad Agr Sci, Fujian Prov Key Lab Genet Engn Agr, Fuzhou, Peoples R China
推荐引用方式 GB/T 7714	Chi, Xu,Zhang, Yingchun,Xue, Zheyong,et al. Discovery of rare mutations in extensively pooled DNA samples using multiple target enrichment[J]. PLANT BIOTECHNOLOGY JOURNAL,2014,12(6):709-717.
APA	Chi, Xu.,Zhang, Yingchun.,Xue, Zheyong.,Feng, Laibao.,Liu, Huaqing.,...&Qi, Xiaoquan.(2014).Discovery of rare mutations in extensively pooled DNA samples using multiple target enrichment.PLANT BIOTECHNOLOGY JOURNAL,12(6),709-717.
MLA	Chi, Xu,et al."Discovery of rare mutations in extensively pooled DNA samples using multiple target enrichment".PLANT BIOTECHNOLOGY JOURNAL 12.6(2014):709-717.

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