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Bt杀虫蛋白基因cry8Ea2的克隆、表达和活性

DOI: 10.7668/hbnxb.2008.04.001, PP. 1-4

Keywords: Bt,cry8Ea2基因,基因克隆,蛋白表达,杀虫活性

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Abstract:

根据已知Btcry8类基因的全长序列设计一对特异性引物JJX5和JJX3,以Bt菌株B-DLL的质粒DNA为模板扩增出3.5kb大小的片段;将该片段插入大肠杆菌表达载体pET-21b中,并完成了该片段的全序列测定。该基因编码区为3495bp,编码的蛋白质由1164个氨基酸残基组成,相对分子质量为131.8kDa,等电点为pH4.71,为弱酸性蛋白。该基因(GenBank:EU047597)编码的氨基酸序列与Cry8Ea1的氨基酸序列同源性高达99.31%,被国际Bt基因命名委员会正式命名为cry8Ea2。经IPTG诱导后该基因在大肠杆菌BL21(DE3)中能够正常表达130kDa的蛋白,表达产物对暗黑鳃金龟、琉璃弧丽金龟和柳蓝叶甲具有活性,在浓度为8.98μg/g时对暗黑鳃金龟、琉璃弧丽金龟一龄幼虫14d的校正死亡率分别为46.67%和55.56%;在浓度为89.8μg/mL时对柳蓝叶甲三龄幼虫96h的校正死亡率为33.33%。

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