棉铃虫触角羧酸酯酶HaCXE6基因的克隆、序列分析与mRNA表达  

Cloning,sequence analysis and mRNA expression levels of the antennal carboxylesterase gene HaCXE6 from Helicoverpa armigera

作　　者：姬继超[1] 王丽莎[1] 赵曼[1] 李为争[1] 原国辉[1] 罗梅浩[1] 郭线茹[1] 

机构地区：[1]河南农业大学植物保护学院,郑州450002

出　　处：《应用昆虫学报》2013年第5期1311-1319,共9页Chinese Journal of Applied Entomology

基　　金：河南省杰出青年科学基金项目(074100510013)

摘　　要：为研究羧酸酯酶(carboxylesterase,COE)在棉铃虫Helicoverpa armigera(Hübner)触角中的生理功能,采用RT-PCR、RACE方法从棉铃虫雄蛾触角中克隆得到一条COE基因全长序列(GenBank No.JX306730),命名为HaCXE6。HaCXE6基因读码框1 623 bp,编码540个氨基酸,预测蛋白质分子量61.02 ku,等电点8.06。氨基酸序列比对分析表明,HaCXE6与海灰翅夜蛾Spodoptera littoralis触角羧酸酯酶CXE6一致性最高,为68%;棉铃虫不同功能羧酸酯酶聚类分析显示HaCXE6与气味降解酶CCE006a最相近。与其它昆虫触角羧酸酯酶多序列比对分析发现,HaCXE6具有酯酶活性必须的催化残基Ser199,Glu323,His434,也保持着α/β-酯酶家族的特征基序Gly-xSer-x-Gly。不同组织和发育时期该基因荧光定量PCR分析结果表明,HaCXE6在雌、雄蛾各个部位均有表达,雌蛾腹部与雄蛾足中表达量最高;卵至成虫各个时期均有表达,蛹中表达量最高。To study the physiological functions of carboxylesterase （ COE ） in Helicoverpa armigera （Hübner）antennae, a new COE gene,named HaCXE6, was cloned from Helicoverpa armigera male antennae （ GenBank No. JX306730 ） by RT-PCR and RACE stratery. The open reading frame is 1 623 bp in length, and encodes 540 amino acid residues with the predicted protein molecular mass of 61. 02 KDa and isoelectric point of 8.06. Multiple sequence alignment analysis revealed that the deduced amino acid sequence has highest identity （ 68% ） with Spodoptera littoralis antennal esterase CXE6. Phylogenetic analysis indicated that HaCXE6 has the closest evolutional relationship with Helicoverpa armigera odorant degrading enzyme CCE006a among different cotton bollworm COEs. The multiple sequence alignment of HaCXE6 and other insect COEs showed that HaCXE6 has the structure characteristic of esterases, including the catalytic triad （ Ser199, Glu323, His434 ） and the conserved mitif Gly-x-Ser-x-Gly in the α/β-esterases family. qPCR was used to analyze the mRNA of HaCXE6 in different tissues and developmental stages in H. armigera. HaCXE6 is a ubiquitous gene and could be detected in all checked tissues and mainly expresses in female abdomen and male legs. And HaCXE6 expresses at all developmental stages and peak in pupa stage.

关 键 词：棉铃虫触角羧酸酯酶 基因克隆 实时荧光定量PCR 功能分析 

分 类 号：S435.622.3[农业科学—农业昆虫与害虫防治]