A high efficiency gene disruption strategy using a positive-negative split selection marker and electroporation for Fusarium oxysporum

doi:10.1016/j.micres.2014.03.004

CORC > 中国农业科学院 > 蔬菜花卉研究所

	A high efficiency gene disruption strategy using a positive-negative split selection marker and electroporation for Fusarium oxysporum
	Liang, Liqin 1,2,3; Li, Jianqiang 2; Cheng, Lin 3; Ling, Jian 1; Luo, Zhongqin 1; Bai, Miao 4; Xie, Bingyan 1
刊名	MICROBIOLOGICAL RESEARCH
	2014
卷号	169 期号:11 页码:835-843
关键词	Split-marker Recombination Homologous Disruption Electroporation Fusarium oxysporum
ISSN号	0944-5013
DOI	10.1016/j.micres.2014.03.004
通讯作者	Liang, Liqin
英文摘要	The Fusarium oxysporum species complex consists of fungal pathogens that cause serial vascular wilt disease on more than 100 cultivated species throughout the world. Gene function analysis is rapidly becoming more and more important as the whole-genome sequences of various F. oxysporum strains are being completed. Gene-disruption techniques are a common molecular tool for studying gene function, yet are often a limiting step in gene function identification. In this study we have developed a F. oxysporum high-efficiency gene-disruption strategy based on split-marker homologous recombination cassettes with dual selection and electroporation transformation. The method was efficiently used to delete three RNA-dependent RNA polymerase (RdRP) genes. The gene-disruption cassettes of three genes can be constructed simultaneously within a short time using this technique. The optimal condition for electroporation is 10 mu F capacitance, 300 Omega resistance, 4kV/cm field strength, with 1 mu g of DNA (gene-disruption cassettes). Under these optimal conditions, we were able to obtain 95 transformants per mu g DNA. And after positive-negative selection, the transformants were efficiently screened by PCR, screening efficiency averaged 85%: 90% (RdRP(1)), 85% (RdRP(2)) and 77% (RdRP(3)). This gene-disruption strategy should pave the way for high throughout genetic analysis in F. oxysporum. (C) 2014 Elsevier GmbH. All rights reserved.
学科主题	Microbiology
语种	英语
出版者	ELSEVIER GMBH, URBAN & FISCHER VERLAG
WOS记录号	WOS:000343378900005
内容类型	期刊论文
源URL	[http://111.203.20.206/handle/2HMLN22E/101857]
专题	蔬菜花卉研究所
作者单位	1.Chinese Acad Agr Sci, Inst Vegetables & Flowers, Beijing 100081, Peoples R China 2.China Agr Univ, Coll Agr Sci & Technol, Beijing 100081, Peoples R China 3.Shanxi Normal Univ, Coll Life Sci, Linfen 041004, Peoples R China 4.Hunan Agr Univ, Coll Hort & Landscape, Changsha 410128, Hunan, Peoples R China
推荐引用方式 GB/T 7714	Liang, Liqin,Li, Jianqiang,Cheng, Lin,et al. A high efficiency gene disruption strategy using a positive-negative split selection marker and electroporation for Fusarium oxysporum[J]. MICROBIOLOGICAL RESEARCH,2014,169(11):835-843.
APA	Liang, Liqin.,Li, Jianqiang.,Cheng, Lin.,Ling, Jian.,Luo, Zhongqin.,...&Xie, Bingyan.(2014).A high efficiency gene disruption strategy using a positive-negative split selection marker and electroporation for Fusarium oxysporum.MICROBIOLOGICAL RESEARCH,169(11),835-843.
MLA	Liang, Liqin,et al."A high efficiency gene disruption strategy using a positive-negative split selection marker and electroporation for Fusarium oxysporum".MICROBIOLOGICAL RESEARCH 169.11(2014):835-843.