关键词: 二化螟, 刷状缘膜囊泡, 苏云金杆菌, Cry1A, 配基印迹

Abstract: Isolation and identification of the binding proteins of Cry1A toxins in the midgut brush border membrane vesicles (BBMV) of Chilo suppressalis is very important for understanding the mode of action and resistance mechanism of Cry1A toxins. In the present study, insecticidal activity and ligand binding of Bacillus thuringiensis Cry1A toxins to the midgut BBMV of striped rice stem borer C. suppresalis were investigated. The results indicated that Cry1Ab was more toxic than Cry1Ac, and Cry1Ac was more toxic than Cry1Aa against the 3rd instar larvae of the laboratory strain (CN) of C. suppresalis. Ligand blot analysis showed that there were six major proteins (50, 70, 90, 120, 160 and 180 kDa) binding to Cry1Ac in the midgut BBMV of the CN strain. Bands of 180, 160 and 90 kDa binding proteins were much darker than the others, suggesting that these proteins had higher binding concentrations. Results of homologous competition binding indicated that the 180 and 90 kDa protein bands were of low binding affinity, while the other four protein bands (160, 120, 70 and 50 kDa) were of high binding affinity. Heterologous competition binding assays were conducted to study crossreactivity of the binding sites between Cry1Ab and Cry1Ac. Cry1Ab competed for all binding sites recognized by Cry1Ac, with high affinity to 180, 120, 70 and 50 kDa proteins and low affinity to 160 and 90 kDa proteins. These data suggest that Cry1Ac and Cry1Ab share several common binding sites in the midgut BBMV of C. suppressalis, but they have different binding affinity with respect to each binding site. Considering the similarity in binding sites, Cry1Ab and Cry1Ac should not be used simultaneously in the transgenic Bt rice to control the target pest C. suppressalis.

Key words: Chilo suppressalis, brush border membrane vesicle (BBMV), Bacillus thuringiensis, Cry1A, ligand blot