中国小麦地方品种抗白粉病基因Pm5e精细遗传图谱绘制

Powdery mildew, caused by Blumeria graminis f. sp. tritici (Bgt), is one of the most important diseases of wheat in China and worldwide. Growing resistant cultivars has proved to be an economical and effective approach to control this disease. Major gene resistance, either alone or stacking together, has extensively employed in wheat breeding programs. Chinese landraces harbored many powdery mildew resistance genes. Among them, Pm5e, derived from Fuzhuang 30, was proved to be resistance to many Chinese and European Bgt isoaltes for decades. A high density genetic linkage map of Pm5e is essential for marker assisted selection (MAS), fine mapping and towarding map-based cloning of the gene. In this proposal, next generation sequencing (NGS) and bulked segregant analysis (BSA) techniques were applied to develop polymorphic SNP markers tightly linked to Pm5e. Combinaing with comparative genomics analysis, a high-density genetic linkage map of Pm5e gene will be constructed using F7:8 recombinant inbred lines and large F2 segregating population of Fuzhuang30/Nongda015. The proposed research will result a fine genetic linkage map of Pm5e which will be severed as framework towarding map-based cloning of the gene. The BSA RNA-Seq methodology applied in this research will demonstrate its potential in developing high density genetic linkage maps of agronomically important traits in common wheat.

白粉病是小麦重要病害，小麦地方品种蕴含丰富的抗白粉病基因。Pm5e是来源于我国小麦地方品种、定位于7BL染色体臂，对国内外白粉病菌系具有良好抗性的抗白粉病基因。构建Pm5e高密度遗传连锁图谱，对于该基因的分子标记辅助选择、精细定位和图位克隆具有重要意义。本项目以复壮30/农大015后代重组自交系(F7:8)和F2代分离大群体为材料，应用下一代转录组测序技术和集群分离分析（BSA）相结合的BSA RNA-Seq方法，开发与小麦抗白粉病基因Pm5e紧密连锁的分子标记，结合比较基因组学分析，构建Pm5e高密度遗传连锁图谱，并在分离大群体中对Pm5e进行精细定位。研究方法和结果对于应用BSA RNA-Seq策略在普通小麦品种间后代群体中进行重要性状的精细遗传作图有重要借鉴和参考意义。

小麦地方品种复壮30携带的抗白粉病基因为Pm5e，多年来对我国各大麦区流行的小麦白粉病菌优势小种和国外的白粉病菌小种表现高抗或免疫，是小麦抗性育种中可贵的抗白粉病资源。在利用SSR标记、比较基因组学等技术均无法找到其紧密连锁分子标记的情况下，我们利用BSR-Seq和比较基因组学的方法开发出29个与Pm5e连锁的分子标记，构建了Pm5e的高密度遗传连锁图谱，开发出与Pm5e共分离的SNP标记WGGC5746、WGGC7051，确定Pm5e所在的7BL染色体WGGC6892-WGGC7051的区段与粗山羊草7DL染色体BM137749-AT7D7135区段BAC contig 3554 (1273.5kb)的同源关系。进而利用Pm5e遗传图谱与粗山羊草scaffold281物理图谱比较定位，将Pm5e候选基因定位在与粗山羊草235 kb区间同源的区间内，该235 kb区间含有1个未知功能的基因，1个丝氨酸蛋白激酶基因和1个R类似基因。目前正在对候选基因验证。.同时，利用标记WGGB7051对405份小麦地方品种材料进行分子标记检测，发现在所有感白粉病材料的WGGB7051位点（C/G）与复壮30的WGGB7051位点（A）均不相同。在23份抗病材料的WGGB7051位点（A）与复壮30的WGGB7051位点（A）一致，表明在这些抗病材料可能含有抗白粉病基因Pm5e。一方面在小麦抗白粉病育种中，标记WGGB7051可以作为辅助选择抗白粉病基因Pm5e的诊断标记，另一方面也表明WGGB7051位点的SNP（A/G）与抗白粉病的功能密切相关。.本项目结果表明：一方面在普通小麦等基因组庞大且复杂的作物或亲本亲缘关系较近的群体中，将比较基因组学和BSR-Seq方法相结合是精细定位基因的有效方法。另一方面可以极大地推进抗白粉病基因Pm5e克隆的进程。