重测序结合转录组测序挖掘东农L-10抗大豆胞囊线虫病3号生理小种基因及其功能鉴定与利用

Soybean cyst nematode (SCN) race 3 was the major race in Chinese main soybean producing areas. Application of the cultivar with resistance to SCN was the effective method to control this disease. A excellent resistant cultivar ‘DongnongL-10’, a susceptible cultivar ‘Heinong37’, and a recombinant intercross line (RIL) population derived from a cross between ‘DongnongL-10’ and ‘Heinong37’, were re-sequencing (Sequencing depth of parents > 22 X, Sequencing depth of population > 3 X). A Bin-Map was constructed, which included 5521 Bins. The total genetic distance of this map is 2103.51cM, the average distance of which was 0.38 cm. Combined with the phenotypic values of the population, a total of 4 QTL with main effect were identified, which could explain more than 10% of phenotypic variations. Also, transcriptome sequencing for a resistance line and a susceptible line have been completed. .Based on these works, ‘DongnongL-10’ with resistance to SCN race 3 would be explored through the combination between re-sequencing and transcriptome sequencing results. These 4 main QTL in ‘DongnongL-10’ were fine-mapping, novel candidate gene of which would be cloning. Function of these new candidate genes would be analyzed via Agrobacterium rhizogenes/Agrobacterium tumefaciens combination with CRISPR/Cas, RNAi, and over expression technology. Functional markers would be developed based on resistance genes. These works would promote SCN resistance breeding process and prepare theory and technology basis of soybean molecular breeding.

大豆胞囊线虫病3号生理小种是我国大豆主产区的优势小种，利用抗病品种是控制该小种的有效措施之一。本项目前期对优异抗源东农L-10和感病品种黑农37及二者杂交衍生的重组自交系进行了重测序（父母本测序深度到22X，群体测序深度3X），构建了1个包括5521个bin 的Bin Map 图谱，其总遗传距离为2103.51cM，平均遗传距离为0.38 cM。结合群体的表型值，获得了4个表型贡献率大于10%的主效QTL。同时，也完成了1对极端抗病和极端感病家系的转录组测序工作。.本项目在此基础上，拟整合重测序和转录组测序结果，从而全面和系统地研究东农L-10对3号生理小种的抗性，精细定位主效QTL并克隆抗病新基因，并通过发根农杆菌结合CRISPR/Cas技术、RNA干涉技术和过量表达技术来快速鉴定抗病候选基因的功能，同时开发实用化标记并验证其准确性，为推动和缩短我国抗SCN育种进程奠定理论和技术基础。

大豆胞囊线虫病3号生理小种是我国大豆主产区的优势小种，利用抗病品种是控制该小种的有效措施之一。传统抗线育种准确性、效率低且遗传进程慢，分子辅助育种可有效克服传统抗线育种的缺点。目前，我国抗线分子辅助育种方面缺乏具有重大利用价值的新基因, 本项目将综合运用重测序、精细定位、转录组测序、分子生物学等手段，挖掘和鉴定优异 优异抗源的新基因，并开发基于抗性基因的功能标记，建立和完善相应的分子 标记辅助育种体系，其对于推动大豆抗线育种具有重要的科学意义。本项目按照研究计划在前期的基础上，获得1张包括5521个bin 的高密度Bin-Map遗传图谱；整合重测序和转录组测序结果，全面和系统地研究东农L-10对大豆胞囊线虫病3号生理小种的抗性，精细定位了4个抗病主效QTL，通过发根农杆菌结合CRISPR/Cas 技术、RNA干涉技术和过量表达技术，来快速鉴定获得4个抗病功能明确的基因，并初步明确抗病基因的作用机理,获得用于筛选抗病种质的功能标记并验证其准确性和效率；利用功能标记辅助筛选获得农艺性状优良且抗病的新材料13份，这在国内外相关研究均鲜见且为推动和缩短我国抗线育种进程奠定理论和技术基础。本项目发表SCI收录论文24篇，国内核心期刊发表论文10篇，申请发明专利3项；通过本课题实施，培养硕士研究生5名，本项目主持人入选国家万人计划科技创新领军人才计划和黑龙江省杰出青年基金计划。