基于基因特异性标记分析Pm21在中国冬小麦品种(系)中的分布  

Distribution of Powdery Mildew Resistance Gene Pm21 in Chinese Winter Wheat Cultivars and Breeding Lines Based on Gene-Specific Marker

作　　者：江峥[1,2] 王琪琳[1,3] 吴建辉[1,3] 薛文波[1,2] 曾庆东[1,3] 黄丽丽[1,3] 康振生[1,3] 韩德俊[1,2] 

机构地区：[1]西北农林科技大学早区作物逆境生物学国家重点实验室,陕西杨凌712100 [2]西北农林科技大学农学院,陕西杨凌712100 [3]西北农林科技大学植物保护学院,陕西杨凌712100

出　　处：《中国农业科学》2014年第11期2078-2087,共10页Scientia Agricultura Sinica

基　　金：国家“973”计划(2013CB127700);国家小麦产业技术体系(CARS-3-1-11);国家公益性行业(农业)科研专项(201203014);高等学校学科创新引智计划项目(B07049)

摘　　要：【目的】来自小麦-簇毛麦6VS/6AL易位系的抗病基因Pm21对小麦白粉病具有持久和广谱抗性,开发该基因的特异性标记,分析其在全国冬麦区中的应用情况,为Pm21的合理布局及分子标记辅助选择育种提供理论依据和技术支撑。【方法】根据已克隆的与Pm21抗性途径紧密相关的丝氨酸/苏氨酸蛋白激酶基因Stpk-V的序列(GenBank登录号为HQ864471.1),提取其蛋白序列并利用Pfam软件分析其保守结构域起止位点,在其保守结构域外设计开发特异序列标记WS-1;构建Pm21载体品种92R137和感病品种Avcoet S(AvS)的F2群体,以小麦白粉病菌E09对该群体每个单株进行苗期抗白粉病表型鉴定,同时利用WS-1对F2群体进行分子检测,分析检测表型与抗病表型,以验证WS-1标记的准确性;利用WS-1标记对来自中国不同冬麦区的662份小麦品种(系)进行分子标记检测,分析Pm21在不同麦区小麦品种(系)的分布情况,并将检测到含有Pm21的品种(系)在田间进行抗白粉病鉴定;选取WS-1已检测到及没有检测到Pm21的品种(系)各50份,利用曹爱忠等开发的标记NAU/xibao15902进行PCR扩增,进一步证明WS-1的准确性。【结果】(1)开发的Pm21特异标记WS-1为显性标记,含Pm21的小麦材料在8%非变性聚丙烯酰胺凝胶中扩增出一条大小为949 bp的片段,而不含该基因的小麦材料中无该片段。(2)在包含377个株系的F2群体中,286个单株为抗病,91个单株为感病,抗感比符合3﹕1的分离比例,表明在该群体中Pm21表现为显性单基因,WS-1对F2群体的每个株系的检测结果与抗/感表型完全一致。(3)供试的662份小麦材料中,49份携带Pm21,平均分布频率为7.4%,其中,西南冬麦区中检测到33份,占该区参鉴品种(系)数的34.4%,而北部冬麦区、黄淮冬麦区和长江中下游冬麦区,分别检测到4份、9份和3份,各占该区参鉴品种(系)数的5.3%、3.1%和1.5%。【结论】开发的Pm21特异性标记WS-1可以作为�Objective Powdery mildew, caused by Blumeria graminis f. sp.tritici （Bgt）, is an important disease that causes substantial yield losses in wheat （Triticum aestivum L.） in China, there is a great significance for using resistance genes to control this disease. Powdery mildew resistance gene Pm21, transferred from Haynaldia villosa by a 6VS/6AL translocation, confers durable and broad-spectrum resistance to the disease. This study is to develop gene specific PCR marker, analyze the distribution of Pm21 in China winter wheat regions, and to provide a theoretical basis and technical support for reasonable arrangement and marker-assisted selection for Pm21.[Method]Based on the sequence of Stpk-V gene which was cloned and related to the resistance pathway （GenBank accession number：HQ864471.1）, the protein sequences were extracted and the conservative start-stop sites were analyzed by Pfam software, in order to amplify this gene specifically, the marker WS-1 was developed excluding conserved domain. For constructing an F2 populations derived from susceptible cultivar Avocet S and the lines 92R137 carrying Pm21, infection types in F2 plants were evaluated by artificial inoculation with isolate E09 during seedling stage, F 2 plants were amplified by WS-1, and the testing results and infection types were analysed to confirm the accuracy of WS-1. A total of 662 wheat cultivars and breeding lines growing in different winter wheat regions in China were detected by WS-1 to analyze the distribution of Pm21, and the materials carrying Pm21 were further tested the resistance to powdery mildew under field conditions. In order to further prove the accuracy of WS-1, the marker NAU/xibao15 902 developed by Cao Aizhong and others was used to amplify the lines with Pm21 and without Pm21 （50 samples from each type）. [Result] As a dominant molecular marker, WS-1 could be amplified a 949 bp fragment in the lines with Pm21 on 8%non-denaturing polyacrylamide gel, but did not in the lines without Pm21. The 377 F2 population

关 键 词：小麦 白粉病 PM21 基因特异性标记 抗病基因持久性 

分 类 号：S512.11[农业科学—作物学]