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苏云金芽孢杆菌BtC008对棉铃虫、小菜蛾和甜菜夜蛾等鳞翅目害虫具有高毒力。本文分析了其杀虫基因类型包括cry1Ab、cry1Ca、cry1Ia、cry2Ab和一种新的cry1类基因,SDS-PAGE分析其至少2种cry1类基因获得了表达,在分子水平说明了该菌株高毒力的原因。在此基础上克隆了1种cry1Ab类杀虫晶体蛋白(insecticidalcrystalproteins,ICPs)基因,并被Bt杀虫晶体蛋白基因命名委员会命名为cry1Ab20。序列分析显示该基因所编码蛋白与已知的Cry1Ab13有1个氨基酸不同。将cry1Ab基因毒性片段(1.95kb)插入表达载体pET-21b,转化EscherichiacoliRosetta(DE3)菌株,表达的包涵体蛋白对小菜蛾和棉铃虫幼虫具有杀虫活性,LC50分别为100.74μg/mL和29.87μg/mL,进一步明确了Cry1Ab20蛋白的杀虫活性区。
Bacillus thuringiensis BtC008 has high virulence against Lepidoptera pests such as cotton bollworm, diamondback moth and beet armyworm. This paper analyzed the insecticidal genotypes including cry1Ab, cry1Ca, cry1Aa, cry2Ab and a novel cry1 gene. SDS-PAGE analysis showed that at least two cry1 genes were expressed, indicating that the strain is highly virulent at the molecular level s reason. Based on this, one cry1Ab insecticidal crystal protein (ICPs) gene was cloned and named cry1Ab20 by the Bt Insecticidal Crystal Protein Gene Nomenclature Committee. Sequence analysis showed that this gene encodes a protein that differs from the known CrylAb13 by 1 amino acid. The cry1Ab genomic fragment (1.95kb) was inserted into expression vector pET-21b and transformed into Escherichia coli Rosetta (DE3) strain. The expressed inclusion protein had insecticidal activity against Plutella xylostella and H. armigera larvae with LC50 values of 100.74μg / mL and 29.87μg / mL, to further clarify the insecticidal activity of Cry1Ab20 protein area.