小菜蛾是首个在田间对Bt毒素产生抗性的世界性害虫，目前已有多项关于抗性机理的报道，但尚缺乏在蛋白质水平上系统揭示抗性机理的研究。本项目拟基于前期已经制备的小菜蛾中肠单克隆抗体组，采用抗体芯片杂交、荧光标记等技术高通量筛选小菜蛾敏感品系(G88)与Cry1Ac抗性品系(抗性倍数>37,037)之间的差异蛋白；采用免疫沉淀、质谱检测、遗传连锁分析等方法鉴定并验证与Bt抗性相关的关键蛋白；采用CRISPR/Cas9、细胞过表达、免疫荧光等技术探明关键蛋白介导Bt抗性的功能，在蛋白质水平上揭示小菜蛾对Bt的抗性机理。本项目是申请人所在团队成功破译小菜蛾基因组以及20多年来从事小菜蛾研究的延续和拓展，为深入研究害虫Bt抗性机理提供了新的思路和方法，对优化害虫抗性治理策略、提高生物农药和Bt作物的控害效能具有重要意义。

The diamondback moth, Plutella xylostella (L.), is the first pest species that evolved resistance to Bacillus thuringiensis (Bt) toxins. Although a number of studies have documented the resistance of P. xylostella to Bt toxins, information on the proteome-based investigation on the resistance mechanism remains limited. The present project intends to explore the resistance mechanisms of P. xylostella to Bt toxin Cry1Ac based on the protein-specific monoclonal antibodies completed recently by our team. To do so, we will screen differentially expressed proteins between a susceptible (G88) and a Cry1Ac-resistant (resistance ratio>37,037) strains using the hybridization of protein-specific antibody arrays and the fluorescence labeling, and then identify and validate the key proteins involved with the Bt resistance based on genetic linkage analyses combined with immunoprecipitation and mass spectrometry detection. Bt resistance-mediated functions of the identified key proteins will be tested and verified using the methods of gene editing (CRISPR/Cas9), cell overexpression, and immunofluorescence, aimed ultimately at uncovering and better understanding the mechanisms of Bt resistance in P. xylostella. This project is a continuation and extension of our previous research on the biology, ecology and genomics of P. xylostella over the past 20 years. Results from this project will bring us novel ideas and methods to further explore the Bt resistance mechanisms of targeted species, optimize the strategies for management of insecticide resistance, and improve the control effectiveness of biological agents and Bt crops.