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抗稻瘟病基因Pik1-H4的克隆及其互作蛋白的筛选

发表时间:2016-08-15  浏览量:1396  下载量:363
全部作者: 董双玉,刘 浩,王 慧,陈志强,王加峰
作者单位: 华南农业大学国家植物航天育种工程技术研究中心
摘 要: 前期的研究推测高抗稻瘟病水稻品系H4对稻瘟病菌(Magnaporthe oryzae)的抗性由Pik位点的基因介导。为进一步确认该基因是否为Pik位点的等位基因并揭示该基因的抗病机理,对该位点基因(Pik1-H4)的编码序列(coding sequence,CDS)进行了克隆,并以Pik1-H4为诱饵,采用酵母双杂交的方法对其互作蛋白进行了筛选。结果表明:Pik1-H4基因的编码序列全长为3.429 kb,与已经报道的Pikh-1相似,仅有两个碱基的突变;利用酵母双杂交系统筛选得到OsBIHD、OsCOL9等6个分别与Pik1-H4具有相互作用的蛋白。进一步对Pik1-H4与OsBIHD、OsCOL9的相互作用利用双分子荧光互补技术(bimolecular fluorescence complementation,BiFC)进行了验证,结果表明Pik1-H4仅能与OsBIHD在细胞核内存在直接的相互作用。本研究不仅克隆了介导H4抗性的基因Pik1-H4,还有助于进一步了解Pik1-H4介导的抗稻瘟病机制。
关 键 词: 作物遗传学;Pik1-H4;酵母双杂交;双分子荧光互补技术;互作蛋白
Title: Cloning of rice blast resistance gene Pik1-H4 and its interaction protein screening
Author: DONG Shuangyu, LIU Hao, WANG Hui, CHEN Zhiqiang, WANG Jiafeng
Organization: National Engineering Research Center of Plant Space Breeding, South China University
Abstract: Previous studies implied that high blast broad spectrum resistance of rice line H4 to Magnaporthe oryzae was mediated by one allele from Pik locus. To further figure out whether the resistance gene is Pik1-H4 and reveal the resistant mechanism of this gene, the coding sequence (CDS) of Pik1-H4 gene was cloned and a yeast two-hybrid screen was performed with a rice leaf cDNA library using Pik1-H4 as a bait. The results showed that the coding sequence of Pik1-H4 is 3.429 kb, which is high similar to that of Pikh-1 with two base-silent mutation. Six interaction proteins, including OsBIHD and OsCOL9 were identified from the yeast two-hybrid screen. Further results showed that Pik1-H4 can interact with OsBIHD but not with OsCOL9 in the cell nucleus in the bimolecular fluorescence complementation (BiFC) assay. The study not only cloned the resistant gene Pik1-H4 but also provided new clues for understanding the mechanism of the Pik1-H4-mediated blast resistance.
Key words: crop genetics; Pik1-H4; yeast two-hybrid; bimolecular fluorescence complementation; interaction protein
发表期数: 2016年8月第15期
引用格式: 董双玉,刘 浩,王 慧,等. 抗稻瘟病基因Pik1-H4的克隆及其互作蛋白的筛选[J]. 中国科技论文在线精品论文,2016,9(15):1521-1529.
 
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