灰飞虱卵巢内水稻条纹病毒增殖机制的研究

Small brown planthopper (SBPH) is the primay vector of rice stripe virus (RSV), which causes the tremendous loss in rice yield. RSV could reproduce in SBPH female ovary and then be transmitted to SBPH progeny via egg, which is the key step of RSV vertical transmission. Unfortunately, it is largely unknown about the molecular mechanism of RSV replication in the SBPH ovary. Currently, the transcriptome of SBPH ovary has been sequenced in our lab.Based on this, we are going to analyze the gene expression profiles of ovary from the female small brown planthoppers with or without RSV infection as well as identify and characterize RSV replication-related genes in ovary using newly developed bioinformatics algorithms combined with real-time PCR amplication. And then RSV replication-related genes and RSV interaction will be studied with GST pull-down technique and western blot. Finally, we knockdown these RSV replication-related genes to answer if these genes affect RSV replication in SBPH ovary using RNAi technique. Taken this together, we are confident to construct signal regulation pathway of RSV replication in ovary and further elucidate the molecular mechanism of RSV replication.

灰飞虱介导水稻条纹病毒（RSV）的循环传播，给水稻生产造成了巨大损失。灰飞虱卵巢是条纹病毒自我增殖的主要场所，是RSV实现介体循环和垂直传播的关键环节，但RSV在灰飞虱卵巢内繁殖的机制仍然不清楚。项目组前期已获得灰飞虱卵巢组织的转录组数据，在此基础上，本项目拟利用新开发的生物信息学算法，分析高带毒种群和不带毒种群间卵巢表达谱的差异，鉴定与条纹病毒在卵内繁殖的关键靶标基因，利用定量PCR技术进一步验证上述相关基因的表达情况。同时通过体外表达获得上述基因的重组蛋白，采用GST pull-down和Western Blot等技术研究这些重组蛋白与条纹病毒蛋白的相互作用。在确定与病毒增殖有关的基因后，通过RNAi技术干扰上述基因的正常表达，鉴定其对RSV病毒在卵巢内增殖的影响。综合上述信息，利用生物信息学分析构建灰飞虱卵巢内条纹病毒增殖的信号调控通路，初步阐述灰飞虱卵巢内条纹病毒的增殖机制。

项目通过研究明确了水稻条纹病毒RSV对于灰飞虱生物学特性尤其是卵巢发育的影响，通过比较转录组学方法分析了带毒与无毒灰飞虱卵巢转录组关键基因的差异表达图谱，克隆并通过RNA干扰方法明确了卵黄原蛋白受体基因（VgR）在RSV病毒增殖过程中的功能，初步明确灰飞虱卵内条纹病毒的增殖机制，这为深入研究RSV病毒与寄主灰飞虱的互作机制奠定基础，将为控制或阻断RSV病毒在灰飞虱体内的传播与增殖途径提供理论指导，为有效防控水稻条纹叶枯病开辟新思路和新手段，解决生产难题。研究结果发表论文4篇，其中SCI 2篇；获发明专利授权1项；培养博士研究生4名、硕士研究生2名。