啶虫丙醚的作用机制研究及其分子靶标的发现

Pyridalyl is a quite effective and low toxic Lepidoptera and Thysanoptera pesticides. But its insecticidal mechanism and target of action are not clear at present. Studies have shown that the inhibitory effect of pyridalyl is highly related to insect ovarian cells. In this project, we would like to get resistant insect populations of diamondback moth and beet armyworm through pyridalyl screening. After screening, ovarian samples of resistant strains and sensitive strains are achieved. The total RNA and proteins of the samples are extracted. The total RNA is sequenced by a new generation of sequencing method (RNA-seq). The differential gene expression between the two populations of diamondback moth and beet armyworm is analyzed by bioinformatics methods. At the same time, the total protein compositions of the resistant and sensitive samples are compared by two-dimensional electrophoresis (2-DE). The significant differential expressed proteins are identified by MALDI-TOF/TOF and LC-MS/MS. Then the monoclonal antibodies of the two strains are prepared. The binding proteins are analyzed by western-blot and mass spectra. Finally, the mechanism and the action target of pyridalyl are proposed by comprehensive comparation of the results of transcriptomics and proteomics. We believe that this project implementation will provide guidance for the rational design of new inhibitors and provide scientific basis for the delay of the resistance development of pyridalyl.

啶虫丙醚是一种针对鳞翅目、缨翅目的高效低毒杀虫剂，其杀虫机理以及作用靶标目前还不明确，研究表明啶虫丙醚的作用与昆虫卵巢细胞的抑制作用高度相关。本项目拟利用啶虫丙醚对小菜蛾以及甜菜夜蛾田间种群进行抗性筛选，得到对啶虫丙醚具有高抗性的昆虫品系，解剖得到抗性以及敏感品系的卵巢样品，提取样品的总核酸RNA以及总蛋白，通过新一代测序方法（RNA-seq）测定抗性以及敏感品系的全转录组，利用生物信息学方法分析测序结果，比较二者的表达差异基因；同时利用双向电泳（2-DE)的方法分析两品系的总蛋白组成，比较分析二者表达差异蛋白，通过MALDI-TOF/TOF和LC-MS/MS鉴定差异表达的蛋白；制备啶虫丙醚的单克隆抗体，结合免疫印迹以及质谱的方法分析啶虫丙醚的结合蛋白；综合比较转录组学以及蛋白质组学的研究结果，提出啶虫丙醚的作用机制以及靶标，为新型抑制剂的合理设计提供指导，为延缓抗性发展提供科学依据。

啶虫丙醚是一种针对鳞翅目、缨翅目的高效低毒杀虫剂，但其杀虫机理以及作用靶标目前还不明确。研究表明啶虫丙醚的作用与昆虫卵巢细胞的抑制作用高度相关。基因组学、蛋白质组学以及生物信息学的研究方法已经广泛应用于医药以及农药作用机制以及靶标的研究。本项目利用啶虫丙醚对小菜蛾以田间种群进行抗性筛选，得到对啶虫丙醚具有高抗性的昆虫品系，解剖得到抗性品系以及敏感品系的卵巢样品，提取样品的总核酸RNA以及总蛋白，通过新一代测序方法（RNA-seq）测定抗性以及敏感样品的全转录组，利用生物信息学方法分析测序结果，比较二者的表达差异基因；同时利用二维电泳的方法（2-DE)分析两品系的总蛋白组成，综合比较转录组学以及蛋白质组学的研究结论，对啶虫丙醚的作用机制以及靶标进行了初步预测.