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分离和鉴定二化螟Chilo suppresalis幼虫中肠刷状缘膜囊泡(BBMV)中Cryl A毒素的受体蛋白,对于阐明Cryl A毒素作用机理和二化螟抗性机理具有十分重要的意义。为此,本文就Cryl A毒素对二化螟杀虫活性及Cryl Ac与二化螟中肠受体的配基结合进行了研究。结果表明:Cryl Ab对二化螟室内品系(CN)的毒力高于Cryl Ac,而Cryl Ac高于Cryl Aa。配基结合分析表明二化螟CN品系幼虫中肠BBMV中有6个Cryl Ac结合蛋白(分子量分别为50,70,90, 120,160和180 kDa),其中180,160和90 kDa结合蛋白的条带颜色明显深于其他结合蛋白的条带,表明这3个受体蛋白具有较高的结合浓度。同源竞争结合研究表明,180和90 kDa结合蛋白为Cryl Ac的低亲合性结合蛋白,其他4个为高亲合性结合蛋白。为了研究Cryl Ac和Cryl Ab受体结合部位的相互作用,进行了异源竞争结合研究。Cryl Ab可以与Cryl Ac所有的6个结合蛋白进行竞争性结合,与180,120,70和50 kDa结合蛋白具有高亲合性,而与160和90 kDa结合蛋白具有低亲合性。结果显示,Cryl Ac与Cryl Ab在二化螟幼虫中肠BBMV上拥有多个共享的结合位点,但对每个结合位点的亲合性有差异。基于毒素结合部位的相似性,Cryl Ac和Cryl Ab不宜同时用于转基因Bt水稻来控制二化螟。
Isolation and identification of the CrylA toxin receptor protein in the midgut brush border membrane vesicles (BBMV) of Chilo suppresalis larvae were important for elucidating the mechanism of CrylA toxin and the mechanism of resistance to S. In this paper, the insecticidal activity of CrylA toxin against Chilo suppressalis and the binding of Cryl Ac to the receptor of Rhizoctonia barramundi were studied. The results showed that the virulence of Cryl Ab to the stem borer (CN) strain was higher than that of Cryl Ac, while that of Cryl Ac was higher than that of Cryl Aa. Ligand binding analysis showed that there were 6 Cryl Ac binding proteins (molecular weights 50, 70, 90, 120, 160 and 180 kDa respectively) in the midgut BBMV of the Chilo suppressalis larvae. The color bands of the 180, 160 and 90 kDa binding proteins were significantly Stronger than the other binding protein bands, indicating that the three receptor proteins have a higher binding concentration. Homologous competition studies show that the 180 and 90 kDa binding proteins are Cryl Ac low affinity binding proteins and the other four are high affinity binding proteins. In order to study the interaction of the Cryl Ac and Cryl Ab receptor binding sites, heterologous competition binding studies were performed. Cryl Ab competes competitively with all six binding proteins of Cryl Ac, with high affinity for 180, 120, 70 and 50 kDa binding proteins and low affinity with 160 and 90 kDa binding proteins. The results showed that Cryl Ac and Cryl Ab had multiple shared binding sites on BBMV of the late-stage Chilo suppressalis larvae, but had different affinity for each binding site. Based on the similarity of toxin binding sites, Cryl Ac and Cryl Ab should not be used simultaneously in transgenic Bt rice to control the rice stem borer.