粘虫颗粒体病毒增效蛋白在苏云金芽胞杆菌中的表达及增效活性  

Expression and synergistic activity of enhancin from Pseudaletia unipuncta granulovirus-Ps in Bacillus thuringiensis

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作  者:黄立鑫 孙俊 刘琴[1] 韩光杰[1] 李传明[1] 夏杨 陆玉荣[1] 徐健[1] HUANG Lixin;SUN Jun;LIU Qin;HAN Guangjie;LI Chuanming;XIA Yang;LU Yurong;XU Jian(National Experimental Station of Agricultural Microbiology in Yangzhou,Lixiahe Institute of Agricultural Sciences,Yangzhou 225007,Jiangsu,China;Yangzhou Luyuan Bio-Chemical Company Limited,Yangzhou 225008,Jiangsu,China)

机构地区:[1]江苏里下河地区农业科学研究所、国家农业微生物扬州观测实验站,江苏扬州225007 [2]扬州绿源生物化工有限公司,江苏扬州225008

出  处:《微生物学报》2023年第4期1460-1471,共12页Acta Microbiologica Sinica

基  金:江苏省农业科技自主创新资金[CX(21)3087,CX(22)1009];江苏省国际合作项目(BZ2020039);扬州市科技项目(现代农业)(YZ2021049)。

摘  要:【目的】在苏云金芽胞杆菌(Bacillus thuringiensis,Bt)中表达截短后的转宿主粘虫颗粒体病毒(Pseudaletia unipuncta granulovirus-Ps,PuGV-Ps)增效蛋白,为构建增效Bt工程菌提供理论基础。【方法】通过对截短后增效蛋白的密码子进行优化,构建增效蛋白及其融合蛋白表达载体,分析不同启动子指导下增效蛋白表达量的变化,明确增效蛋白对Bt的增效活性。【结果】本研究构建了表达载体pHTPcry1AcCoEn81、pHTRHCoEn81和pHTNCCoEn81,SDS-PAGE结果显示pHTPcry1AcCoEn81和pHTNCCoEn81分别可以产生81 kDa和134 kDa的重组蛋白。启动子Pcry1Ac和Pcry8E指导下的增效蛋白表达量和重组增效蛋白产量均无显著性差异。生物测定结果表明,重组增效蛋白可以显著增加Bt对小菜蛾的杀虫活性。【结论】研究结果表明,密码子优化的PuGV-Ps增效蛋白可以在Bt中表达并具有显著增效活性,为高效苏云金芽胞杆菌工程菌的构建及应用提供理论指导。[Objective]To express the truncated fragments of enhancin gene from Pseudaletia unipuncta granulovirus-Ps(PuGV-Ps)in Bacillus thuringiensis(Bt)and provide a theoretical basis for the construction of Bt engineering bacteria.[Methods]The codon of the truncated fragments of enhancin gene was optimized for the construction of the expression vectors of enhancin and the expression of fusion proteins.Then,the expression levels of enhancin under the guidance of two promoters were analyzed,and the synergistic activity of enhancin on Bt was determined.[Results]The expression vectors pHTPcry1AcCoEn81,pHTRHCoEn81,and pHTNCCoEn81 were constructed in the study.SDS-PAGE showed that pHTPcry1AcCoEn81 and pHTNCCoEn81 produced recombinant proteins of 81 kDa and 134 kDa,respectively.There was no significant difference in the expression level of enhancin or the yield of recombinant protein under the guidance of promoters Pcry1Ac and Pcry8E.Furthermore,the recombinant enhancin significantly increased the insecticidal activity of Bt against Plutella xylostella.[Conclusion]The codon-optimized enhancin of PuGV-Ps can be expressed in Bt and has significant synergistic activity,which provides theoretical guidance for the construction and application of Bt engineering bacteria with high efficiency.

关 键 词:粘虫颗粒体病毒 苏云金芽胞杆菌 增效蛋白 增效活性 

分 类 号:S476.1[农业科学—农业昆虫与害虫防治]

 

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