大豆抗疫病基因RpsZS18的功能鉴定及抗病机制解析

Phytophthora root rot (PRR) caused by Phytophthora sojae is a destructive disease of soybean. The deployment of resistant cultivar is the most effective strategy to control this disease. However, new race of P. sojae is constantly emerging, resulting in resistance loss easily. Thus, novel Rps gene discovery is particularly important in order to sustainably and effectively control the disease. In our previous studies, a novel Rps gene (RpsZS18) was identified in soybean Zaoshu18 with broad-spectrum resistance to P. sojae, which was mapped in a 112.6 kb interval region on chromosome (Chr.) 2 without typical R gene. RpsZS18 is the only one Rps gene on Chr.2 to date, indicating that it might be a novel type R gene, differing from the known Rps gene. This study will take a larger segregated population and re-sequencing data of multiple cultivars as materials, which are used to narrow the RpsZS18 target region by developing the new markers. The best candidate gene (s) will be confirmed using the bioinformatics and gene expression analysis, whose haplotype diversity and functional markers will be exploited. The function and characteristics of RpsZS18 gene will revealed by functional complementation, spatio-temporal expression, sub-cellular localization experiments. At the same time, RpsZS18 mechanism resistance to P. sojae in Zaoshu18 will be revealed using comparative transcriptomics analyses. This study is expected to identify a new type Rps gene, develop validate functional marker and reveal the new mechanism of soybean resistance to P. sojae, which will lay a foundation for rapidly pyramiding multiple Rps gene to develop broad-spectrum and durable soybean varieties resistance P. sojae.

大豆疫病是由大豆疫霉引起的毁灭性病害，利用抗病品种是防治该病害最有效的途径。但由于大豆疫霉新小种不断出现，使其抗性极易丧失，为持续有效防治该病害不断挖掘新抗病基因尤为重要。项目组前期在广谱抗疫病大豆早熟18中鉴定一个抗疫病新基因RpsZS18，位于2号染色体无典型抗病基因的112.6kb区域，RpsZS18是此染色体上迄今唯一的Rps基因，提示可能是一个不同于已知Rps基因的新的抗病基因类型。本项目拟以大群体和多品种重测序数据为材料，缩小RpsZS18目标区域；利用生物信息学及基因表达分析确定最佳候选基因并鉴定其单倍型多样性，开发功能标记；通过功能互补、时空表达、亚细胞定位等研究明确RpsZS18的功能及特征。同时通过比较转录组解析大豆抗疫病调控机制。本项目有望鉴定大豆抗疫病基因新类型，揭示大豆对疫病抗性新机制，获得功能标记，为快速多基因聚合育种培育广谱、持久大豆抗疫病新品种奠定基础。

大豆疫病（又称大豆疫霉根腐病）是由大豆疫霉菌(Phytophthora sojae)引起的限制大豆生产的重要病害之一。挖掘和利用大豆抗疫病基因（Rps）是防治该病害的最有效途径。目前已经鉴定的抗疫病基因Rps大部分位于少数几条染色体的抗病基因富集区域，被初步鉴定或预测为典型抗病基因。项目组前期在大豆早熟18的2号染色体上首次鉴定了唯一一个Rps基因RpsZS18。本项目以早熟18和Williams构建的遗传群体和含有已知Rps基因的大豆品种为材料，通过群体和全基因组重测序数据，将RpsZS18目标区域缩小至71.3kb的物理区间。在RpsZS18候选区间内开发获得了3个共分离的分子标记ZCSSR33、ZCSSR46和ZCindelzs18。基于大豆品种全基因组重测序数据，鉴定出RpsZS18候选区间含有447个SNP和141个InDel，并且有6个基因具有早熟18的特异的单倍型。两个是具有延伸因子的钙结合蛋白基因；一个是编码糖激酶的基因；一个是具有亮氨酸拉链的丝氨酸/苏氨酸蛋白激酶（STK）基因，另外两个为无功能注释的基因。通过对6个基因进行时空和组织的差异表达分析，发现在根部、下胚轴叶片中，仅有3个基因检测到表达，并在抗病亲本中的表达量均高于感病亲本。被预测为RpsZS18的候选基因。通过抗感亲本接种后的转录组数据在不同时间点的相关性、主成分、差异基因表达、Go和KEGG分析，发现在接种后48h是抗病转录调控的关键点，48h时与植物抗性相关的主要通路的基因出现明显差异。通过转录组分析显示一个基因各时间点在感病品种Williams中均不表达，而在抗病品种早熟18中均显著表达。推测该基因是一个最佳的候选基因并且是长链非编码的基因，是一个新的大豆抗疫病基因类型，已获得了转基因植株。本项目中开发的分子标记可加促大豆抗疫病育种进程，鉴定的RpsZS18为一个大豆抗疫病基因新类型，为揭示大豆对疫病抗性的新机制，为快速多基因聚合育种培育广谱、持久大豆抗疫病新品种奠定基础。发表相关SCI 学术论文2篇，获得国家发明专利授权一项。