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当前位置: 首页 > 文章 > 斜纹夜蛾保幼激素环氧水解酶基因的克隆和原核表达 河南农业科学 2018 (12) 84-89
Position: Home > Articles > Cloning and Prokaryotic Expression of JHEH Gene from Spodoptera litura Journal of Henan Agricultural Sciences 2018 (12) 84-89

斜纹夜蛾保幼激素环氧水解酶基因的克隆和原核表达

作  者：
张丽丽;武怡琼;杨正飞;郭丽;魏佳平;阚云超
单  位：
南阳师范学院/河南省伏牛山昆虫生物学实验室;信阳师范学院生命科学学院/大别山农业生物资源保护与利用研究院
关键词：
斜纹夜蛾;保幼激素环氧水解酶;克隆;序列分析;原核表达;重组蛋白
摘  要：
为分析斜纹夜蛾(Spodoptera litura)保幼激素环氧水解酶(Juvenile hormone epoxide hydrolase,JHEH)基因在其生长发育过程中的功能,采用反转录聚合酶链式反应(RT-PCR)扩增SlJHEH基因的开放阅读框,并进行原核表达。SlJHEH基因开放阅读框为1 389 bp,编码462个氨基酸,预测蛋白质分子质量为52 ku,等电点为8. 68。氨基酸序列存在催化三联体Asp226、Glu402和His429氨基酸残基及阴氧离子洞Tyr297、Tyr372和HGWP花样结构氨基酸残基。系统发育树分析结果表明,Sl JHEH与棉铃虫(Helicoverpa armigera) JHEH亲缘关系最近,氨基酸序列同源性高达78%。将SlJHEH的编码区连接到原核表达载体p ET32a上,成功构建了重组载体pET32aSlJHEH,转入大肠杆菌BL21(DE3)感受态细胞中进行诱导表达。对表达产物进行SDS-PAGE和Western blotting检测,结果显示,斜纹夜蛾JHEH基因在大肠杆菌中得到高效且准确的表达。
译  名：
Cloning and Prokaryotic Expression of JHEH Gene from Spodoptera litura
作  者：
ZHANG Lili;WU Yiqiong;YANG Zhengfei;GUO Li;WEI Jiaping;KAN Yunchao;College of Life Sciences/Institute for Conversation and Utilization of Dabie Mountains Agrobioresource,Xinyang Normal University;Nanyang Normal University/Henan Key Laboratory of Insect Biology in Funiu Mountain;
单  位：
ZHANG Lili%WU Yiqiong%YANG Zhengfei%GUO Li%WEI Jiaping%KAN Yunchao%College of Life Sciences/Institute for Conversation and Utilization of Dabie Mountains Agrobioresource,Xinyang Normal University%Nanyang Normal University/Henan Key Laboratory of Insect Biology in Funiu Mountain
关键词：
Spodoptera litura;;Juvenile hormone epoxide hydrolase;;Cloning;;Sequence analysis;;Prokaryotic expression;;Recombinant protein
摘  要：
In order to investigate the function of juvenile hormone epoxide hydrolase( JHEH) gene in growth and development of Spodoptera litura,the open reading frame of SlJHEH was cloned by reverse transcription-polymerase chain reaction( RT-PCR) and expressed prokaryotically. The open reading frame was 1 389 bp in length,and encoded 462 amino acids. Its predicted molecular mass and isoelectric point were 52 ku and 8. 68,respectively. The residues Asp226,Glu402,His429 that formed the potential catalytic triad,and the residues Tyr297,Tyr372,and the HGWP motif that constituted the potential oxyanion hole were found in the deduced amino acid sequences. Sl JHEH had a closely phylogentic relationship with JHEH of Helicoverpa armigera,with the amino acid sequence identity reaching 78%. The recombinant p ET32a-SlJHEH was constructed by ligating the code sequence of SlJHEH with prokaryotic expression vector p ET32 a. Then the recombinant genetic engineering bacteria to express JHEH were obtained by transforming pET32a-Sl JHEH into competent cells of Escherichia coli BL21( DE3). The results of SDS-PAGE and Western blotting analysis showed that the expression of Sl JHEH gene in E. coli was high and accurate,which provided the promise for functional study of SlJHEH gene.

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