Bt Cry2Ab对美国白蛾的杀虫活性及与几种中肠受体蛋白体外结合特性分析  

Insecticidal Activity of Bt Cry2Ab on Hyphantria cunea and Its Binding Characteristics with Some Midgut Receptor Proteins

作　　者：王哲 赵丹[1] 常梦颖 赵坤莉 乔英翠 陆秀君[1,2] 郭巍 Wang Zhe;Zhao Dan;Chang Mengying;Zhao Kunli;Qiao Yingcui;Lu Xiujun;Guo Wei(College of Plant Protection,Agricultural University of Hebei,Baoding Hebei 071001,China;Research Center of Wal-nut of Hebei,Baoding Hebei 071001,China)

机构地区：[1]河北农业大学植物保护学院,河北保定071001 [2]河北省核桃工程技术研究中心,河北保定071001

出　　处：《蚕业科学》2020年第4期435-440,共6页ACTA SERICOLOGICA SINICA

基　　金：河北省重点研发计划项目(20326812D);国家重点研发计划项目(2018YFD0201000);现代农业产业技术体系建设专项(CARS-13);河北省自然科学基金项目(C2017204072)。

摘　　要：美国白蛾(Hyphantria cunea)是危害我国蚕区桑树的重要害虫之一,为明确苏云金杆菌(Bacillus thuringiensis,Bt)杀虫晶体蛋白对其作用机制,本文利用大肠埃希菌表达苏云金杆菌Bt Cry2Ab蛋白,在活性测定基础上进行毒素蛋白与美国白蛾中肠类钙黏蛋白(cadherin)功能区(HcCAD-1、HcCAD-2)、碱性磷酸酶(alkaline phosphatase)HcALP1、HcALP3的结合特性分析。原核表达并纯化的Bt Cry2Ab蛋白用美国白蛾3龄幼虫进行生物测定,对幼虫生长发育抑制作用明显,当质量浓度为224 mg/L时抑制率为71.14%,抑制中浓度为68.44 mg/L。将Bt Cry2Ab毒素分别与美国白蛾HcCAD-1、HcCAD-2、HcALP1、HcALP3重组蛋白进行体外配体印迹试验,Bt Cry2Ab与HcCAD-2、HcALP1重组蛋白发生特异结合,而与HcCAD-1、HcALP3不结合,推测HcCAD-2、HcALP1可能为美国白蛾Bt Cry2Ab蛋白的中肠受体蛋白。Hyphantria cunea is one of the main pests of mulberry tree in sericulture area in China. In order to clarify functional mechanism of Bacillus thuringiensis(Bt) insecticidal crystal proteins on Hyphantria cunea, Bt Cry2 Ab protein was expressed by Escherichia coli, and its ligand binding properties with functional areas of cadherin(HcCAD-1, HcCAD-2) and alkaline phosphatase(HcALP1, HcALP3) from the midgut of H. cunea were analyzed based on activity determination in this study. Then, Bt Cry2 Ab protein expressed and purified from prokaryote was bioassayed by H. cunea of the 3 rd instar, which showed significant inhibition on the growth and development of H. cunea. When the concentration of Bt Cry2 Ab protein was 224 mg/L, inhibition rate was 71.14% and median inhibitory concentration was 68.44 mg/L. According to in vitro ligand blot of Bt Cry2 Ab with HcCAD-1, HcCAD-2, HcALP1 and HcALP3 recombinant protein, Bt Cry2 Ab could specifically bind to HcCAD-2 and HcALP1, while not to HcCAD-1 and HcALP3. It is speculated that HcCAD-2 and HcALP1 might be the midgut receptors of Bt Cry2 Ab.

关 键 词：美国白蛾 Bt Cry2Ab蛋白 类钙黏蛋白 碱性磷酸酶 配体印迹 

分 类 号：S888.739[农业科学—特种经济动物饲养] TQ458[农业科学—畜牧兽医]