利用改良SOE-PCR技术定点突变TuMVC4-VPg基因  

Site-directed Mutagenesis of Tu MV C4-VPg Gene by Optimized Splicing by Overlap Extension PCR

作　　者：李国亮[1] 钱伟[1] 张淑江[1] 李菲[1] 章时蕃[1] 张慧[1] 谢露露[1] 武剑[1] 王晓武[1] 孙日飞[1] LI Guo-liang;QIAN Wei;ZHANG Shu-jiang;LI Fei;ZHANG Shi-fan;ZHANG Hui;XIE Lu-lu;WU Jian;WANG Xiao-wu;SUN Ri-fei(Institute of Vegetables and Flowers,Chinese Academy of Agricultural Sciences,Beijing 100081,China)

机构地区：[1]中国农业科学院蔬菜花卉研究所,北京100081

出　　处：《中国蔬菜》2017年第10期39-44,共6页China Vegetables

基　　金：国家自然科学基金项目(31372069);"十二五"国家科技支撑计划项目(2013BAD01B04-03);农业部园艺作物生物学与种质创制重点实验室项目

摘　　要：芜菁花叶病毒(Turnip mosaic virus,Tu MV)是白菜类蔬菜生产上的重要病害,其中Tu MV VPg基因在Tu MV侵染白菜类蔬菜的过程中起着至关重要的作用,且不同Tu MV株系的致病性不同。本试验采用改良的重叠延伸PCR(SOE-PCR)技术,以Tu MV C4-VPg基因序列为模板,定点突变Tu MV C4-VPg与Tu MV CDN1-VPg基因间5个差异核苷酸位点(决定5个氨基酸差异)。结果表明:通过SOE-PCR技术获得了5个Tu MV C4-VPg基因的单点突变体,即Tu MV C4-VPg-Ⅰ、Tu MV C4-VPg-Ⅱ、Tu MV C4-VPg-Ⅲ、Tu MV C4-VPg-Ⅳ和Tu MV C4-VPg-Ⅴ。Turnip mosaic virus(TuMV)was an important disease for Brassica rapa L.ssp.pekinensisproduction,and the TuMV VPg gene had played a vital role infecting production process.While,thepathogenicity of different TuMV strains was diverse.This experiment adopted improved splicing by overlapextension PCR(SOE-PCR)technology,took TuMV-C4VPg gene sequence as template,carried out sitedirectedmutagenesis on5different single-nucleotides between TuMV C4-VPg and TuMV CDN1-VPg genes.The results indicated that5single point mutants of TuMV C4-VPg were obtained by the optimized SOE-PCR,i.e.TuMV C4-VPg-Ⅰ,TuMV C4-VPg-Ⅱ,TuMV C4-VPg-Ⅲ,TuMV C4-VPg-Ⅳand TuMV C4-VPg-Ⅴ.

关 键 词：白菜类蔬菜 芜菁花叶病毒 VPg基因 重叠延伸PCR技术 单点突变 

分 类 号：S436.34[农业科学—农业昆虫与害虫防治]