棉铃虫钙粘蛋白n 端多肽多克隆抗体制备及对bt 抗性 - 应用昆虫学报.pdf

昆虫知识 Chinese Bulletin of Entomology 20 10 ，47 (2 ):293 ～ 298 N 棉铃虫钙粘蛋白 端多肽多克隆抗体制备及 * Bt 对 抗性的初步检测 1  1 2 1  2 黄晶晶 韩 斌 常菊花 沈文飚 沈晋良 (1. 南京农业大学生命科学学院 南京 2 10095 ;2. 南京农业大学植物保护学院 南京 2 10095 ) Preparation of polyclonal antibody of N -terminal peptide of cadherin of H elicoverp a armig era and 1  1 2 primary detection of Bt-resistance. HUANG Jing-Jing ，HAN Bin ，CHANG Ju-Hua ，SHEN Wen- 1  2 Biao ，SHEN Jin-Liang (1. College of Lif e Science ，Nanj ing Agricultural University ，Nanjing 2 10095 ， China ;2. College of Plant Protection ，Nanj ing Agricultural University ，Nanjing 2 10095 ，China) Abstract Rabbit polyclonal antibodies were prepared with recombinant N-terminal peptide of Helicoverp a armigera (Hübner)cadherin ，and were used for identification of Bt-resistance . N-terminal peptide fragment encoding gene Cad285 was amplified by RT -PCR from midgut of H . armigera and cloned to prokaryotic expression vector pET -30 a ，then induced expressing in E. coli BL2 1 with IPTG . Recombinant protein (～ 35 ku)was highly expressed and existed as inclusion bodies. The inclusion bodies were purified with denaturing ，purifying by Ni-NTA and refolding . The purified recombinant Cad285 was used to immunize rabbit for preparing polyclonal antibody with higher titer than 1∶ 16 000 measured by ELISA . Finally ，a laboratory strains of H . armigera was detected by western blot with the polyclonal antibody . The results showed a significant