关键词: 苏云金杆菌以色列亚种, CryIVD基因, PCR, 基因表达, 蚊虫

Abstract: 　Objective To clone and express Bacillus thuringiensis subsp. isrealensis(B.t.i.) crystal protein CryIVD gene and determine its mosquito larvicidal activity. Methods The gene encoding CryIVD (2.0 kb or so) was amplified by PCR, the amplified fragment was inserted into E.coli plasmid pUC18 to construct the recombinant cloning and expression vector pUC18 CryIVD, which was named pUC18 1. The ligation was transformed into competent E.coli DH 5α and the recombinant vector pUC18 1 was confirmed by restriction enzyme digestion and DNA sequencing. After being inducted by IPTG, the expression of CryIVD gene in positive clone was detected by SDS PAGE and the mosquito larvicidal activity of CryIVD was also determined by standard bioassay. Results The results showed that the CryIVD gene was successfully cloned and expressed in E.coli DH 5α . Mosquito larvicidal activity of engineered E.coli (LC 50 ) to Cx.pipiens pallens and Ae.albopictus Ⅱ-Ⅲ instar larvae was 2.38×10 6 cells/ml and 1.6×10 7cells/ml respectively. Conclusion The CryIVD gene was successfully cloned and expressed, and a high mosquito larvicidal activity was observed.

Key words: Bacillus thuringiensis subsp. israelensis, CryIVD gene, PCR, gene expression, mosquito