甜菜夜蛾几丁质脱乙酰酶抵御病毒侵染的分子机制研究

项目名称： 甜菜夜蛾几丁质脱乙酰酶抵御病毒侵染的分子机制研究

项目编号： No.31471775

项目类型： 面上项目

立项/批准年度： 2015

项目学科： 农业科学

项目作者： 郭巍

作者单位： 北京农学院

项目金额： 82万元

中文摘要： 昆虫中肠表面覆盖有无脊椎动物所特有的几丁质-蛋白质的复合结构，围食膜（peritrophic membrane,PM），具有保护中肠免受病毒等微生物侵染等功能，几丁质脱乙酰酶（chitin deacetylase,CDA）是昆虫PM的重要蛋白组分。本项目以农业上重要害虫甜菜夜蛾为研究对象，利用ITRAQ等蛋白质组学方法，结合前期得到的甜菜夜蛾中肠cDNA表达文库ESTs，进行CDA蛋白的分离鉴定及其cDNA基因克隆；利用生物信息学分析其序列结构特性，进行组织特异性表达和生化特性分析；结合转录组测序和表达谱分析，明确病毒侵染后CDA的表达变化，分析病毒侵染后PM通透性的变化，以论证中肠特异性表达的CDA蛋白在病毒侵染后的下调导致围食膜通透性下降而降低了甜菜夜蛾对病毒的敏感性的这一分子机制假说，即CDA蛋白为生防因子病毒的作用靶标，探寻以环境友好型的新手段防治和控制甜菜夜蛾危害。

中文关键词： 甜菜夜蛾；基因克隆；功能分析；分子生物学；基因功能

英文摘要： The peritrophic membrane(PM)is a chitin and glycoprotein layer lines the inbertebrate midgut，which has important roles in facilitating food digestion and providing protection to the midgut epithelium from pathogens infectious peros. Chitin deacetylase(CDA) is one of the major constituents of the PM proteins. This research is centered on the understanding of the biochemistry and molecular biology of one of the key insect pests in China-Spodoptera exigua (S.exigua) midgut CDA. Proteomics and genomics approach will be used to identify and characterize CDA from S.exigua larvae.The cDNA clones code for CDAs will be subjected to sequencing after restriction enzyme digestion analysis to exclude identical clones. The chitin-binding activity of CDAs will be analyzed using the chitin-binding assaying method. Recombinant baculoviruses will be constructed using the Bac-to-Bac system to over-express and analyze the temporal expression of CDAs. Combined transcriptome sequencing with gene expression profile, the downregulation of CDA after pathogen(virus) infection is hypothesized that this protein might be responsible for the homeostasis of the PM structure. The insect may reduce PM permeability by reduction of CDA expression, as a result, decreasing the entrance of baculovirus. These will support the downregulation of CDA as a possible mechanism used by S.exigua to reduce susceptibility to baculovirus by decreasing PM permeability. The mechanism that CDA is the target substrate for baculovirus will be used to explore novel bioagents for pest control.

英文关键词： Spodoptera exiguae;gene clone;functional analysis;molecular biology;gene function