高抗白粉病小麦-两芒山羊草2Mb易位系的创制及抗病相关基因发掘

Powdery mildew is one of the most widespread diseases of wheat worldwide. Breeding resistant cultivars by utilization of powdery mildew resistance genes is considered to be the most economical and effective method of controlling this disease. It was found that Aegilops biuncialis chromosome 2Mb carried novel powdery mildew resistance gene(s) in our previous research. In this study, wheat-Ae. biuncialis 2Mb disomic addition line TA7733 will be used as bridge material to produce wheat-Ae. biuncialis 2Mb translocation lines by crossing the Chinese Spring (CS) ph1b mutant, which significantly elevates the frequency of homoeologous chromosome pairing and homoeologous recombination. The translocation lines will then be characterized by in situ hybridization (ISH) and molecular marker analysis, and evaluated for powdery mildew resistance at seedling stage in the greenhouse and at adult stage in the fields to select translocation lines with small 2Mb segments carrying high resistance gene(s) to powdery mildew. The novel gene(s) conferring powdery mildew resistance will be further cytologically mapped to the corresponding region of chromosome 2Mb, and the closely linked molecular markers will be developed. Furthermore, transcriptome sequencing will be applied to obtain extensive information about the transcriptomes of the disease-resistant addition lines TA7733 and susceptible recipient parent CS after inoculating Bgt isolates. Subsequently, differential expression genes between different materials will be analyzed to discover genes related to powdery mildew resistance locating on 2Mb chromosome by combining chromosomal localization of resistance gene(s). This study will not only provide novel powdery mildew resistance gene(s) and co-segregated molecular markers for wheat disease breeding, but also lay solid foundations for identifying resistance-related genes and further dissecting molecular and genetic mechanisms of disease resistance.

小麦白粉病是一种世界性小麦病害，发掘抗性基因和培育抗病品种是减少白粉病损失最经济有效的途径。本课题组前期研究发现，两芒山羊草2Mb染色体携带白粉病抗性新基因。本项目拟以小麦-两芒山羊草2Mb二体附加系TA7733为材料，利用中国春ph1b基因诱导部分同源染色体之间高频重组，创制小麦-两芒山羊草2Mb易位系，并对易位系进行原位杂交鉴定、分子标记分析和抗性鉴定，筛选高抗白粉病的2Mb小片段易位系，定位抗性新基因于2Mb染色体特定区段，并获得与该基因紧密连锁的分子标记。对接种白粉菌前后的2Mb附加系和受体亲本中国春进行转录组测序和差异表达基因分析，并结合抗病基因区段定位结果，发掘位于2Mb染色体上的抗病相关基因。该项目的实施不仅为小麦白粉病抗性育种提供抗病新基因和共分离的分子标记，还为克隆该抗病基因以及进一步解析抗病分子机理奠定基础。

小麦白粉病是严重影响小麦产量的主要病害之一，发掘抗病新基因是减少白粉病损失最经济有效的途径。本研究前期发现小麦-两芒山羊草2Mb二体附加系TA7733抗白粉病且抗病新基因（Pm2Mb）来自两芒山羊草2Mb染色体。为了有效利用两芒山羊草2Mb染色体上的抗白粉病新基因，本项目：（1）根据小麦-两芒山羊草2Mb二体附加系TA7733转录组序列和小麦全长cDNA序列设计引物，开发了62个两芒山羊草2Mb染色体特异分子标记；（2）利用中国春ph1b突变体与小麦-两芒山羊草2Mb二体附加系TA7733杂交，创制了ph1b纯合且携带2Mb染色体的BC1F2代分离群体，利用4个2Mb染色体长臂和短臂特异分子标记从1000个单株中筛选到12种不同类型的65个两芒山羊草2Mb易位系；经过GISH和FISH鉴定，其中5个为2Mb大片段易位、6个为小片段易位、1个为中间插入易位；（3）对12种不同类型的65个2Mb易位系家系进行抗病性鉴定和分子标记分析，将抗白粉病新基因Pm2Mb定位于2Mb染色体长臂的FL 0.49-0.66区间。同时，获得2个抗病2Mb小片段易位系以及6个2Mb抗病新基因紧密连锁标记；（4）利用抗病小麦-两芒山羊草2Mb易位系构建了F2次级分离群体，经过分子标记和白粉病抗性鉴定，将Pm2Mb基因进一步定位在分子标记2SL-8和CL75697之间，相对小麦2D染色体566.8-591.2 Mb（24.4 Mb）区间。结合Pm2Mb精细定位结果，采用转录组测序和共线性分析，共挖掘到52个两芒山羊草2Mb染色体白粉病抗性相关基因。此外，发表4篇SCI论文，培养研究生3名。本项目为小麦抗病育种提供了抗白粉病新种质及配套分子标记辅助选择技术，为进一步克隆该抗病基因以及解析小麦抗白粉病分子机理奠定了基础。