应用real-time PCR定量检测田间小麦条锈菌潜伏侵染的研究  

Quantification of latent infection of wheat stripe rust in the fields using real-time PCR

作　　者：闫佳会[1] 骆勇[1,2] 潘娟娟[1] 王海光[1] 金社林[3] 曹世勤[3] 马占鸿[1] 

机构地区：[1]中国农业大学植物病理学系农业部植物病理学重点开放实验室,北京100193 [2]美国加州大学Kearney农业研究中心,parlierca93648 [3]甘肃省农业科学院植物保护研究所,兰州730070

出　　处：《植物病理学报》2011年第6期618-625,共8页Acta Phytopathologica Sinica

基　　金：公益性行业(农业)科研专项(200903035;200903004);国家自然科学基金项目(31071642)

摘　　要：为快速、及时地定量检测处于潜育状态下的小麦条锈菌菌量,本研究于2009-2010年实施田间试验,在北京和甘肃各设3个试验小区,将小区分割成1 m×1 m或1.5 m×1.5 m的小格。初春刚返青叶片出现症状前,在每小格内采集30片叶,提取叶片基因组DNA,应用已建立的real-time PCR定量测定方法,计算每个样本的分子病情指数(molecular-detected disease index,MDI)。在本试验的天气条件和试验设计下,采样10 d后,调查田间标记采样点的发病情况,计算病情指数DI,比较MDI和DI的关系,建立回归模型。结果表明:甘肃甘谷3个试验小区的MDI和DI存在极显著的相关性(R12=0.8136,R2=0.8845,R32=0.5928);北京上庄3个试验小区的MDI和DI也存在极显著的相关性(R21=0.7423,R2=0.5784,R32=0.8584)。本研究证明了应用分子生物学方法可估测小麦条锈病潜育侵染程度的可能性,为建立分子定量测定田间潜育侵染量系统提供了可靠依据。Wheat stripe rust,caused by Puccinia striiformis f.sp.tritici（Pst）,is one of the most important wheat disease world wide.The key issue of the disease forecast is to timely and accurately quantify the latent infection levels in overwintering and oversummering volunteer seedlings that may serve as sources of initial inoculum of epidemics.The field experiments were conducted with nature infections in Gangu（Gansu Pro-vince）,and with artificial inoculations in Shangzhuang,Beijing from 2009 to 2010.There were 3 test plots each in Beijing and Gansu,each plot was divided into 36 grids with same area.In early spring,the new appeared leaves were collected for DNA extraction.The real-time PCR assay was applied to quantify both host DNA and the Pst DNA from each sample.The proportion of Pst DNA（pg） over the total amount of wheat DNA（ng） was assigned as the molecular-detected disease index（MDI）.The visual disease index（DI） in each plot was recorded 10 days after leaf sampling.Linear regression between MDI and DI was performed using the corresponding data set.The regressions were significant for both Gangu fields（R21=0.813 6,R22=0.884 5,R23=0.592 8）and Shangzhuang fields（R21=0.742 3,R22=0.578 4,R23=0.858 4）.This study demonstrated the potential applications of real-time PCR in quantification of latent infection level for various research objectives.

关 键 词：PUCCINIA STRIIFORMIS 分子流行学 

分 类 号：S511[农业科学—作物学]