过氧化氢酶(catalase,CAT)作为生物体内的重要物质,其主要功能是参与活性氧代谢过程,在清除H2O2、超氧自由基和过氧化物以及阻止羟基自由基形成等方面发挥着重要作用。本研究利用RT-PCR技术和RACE方法首次克隆和分析了甜菜夜蛾Spodoptera exigua(Hübner)CAT基因,命名为SexiCAT,GenBank登录号为JN051294,其cNDA序列全长为1755bp,开放阅读框长1524bp,推测编码507个氨基酸。经氨基酸序列比对,此多肽序列具有高度保守性,与其他昆虫CAT的序列一致性分别为:家蚕Bombyxmori(87%)、黑腹果蝇Drosophila melanogaster(73%)、埃及伊蚊Aedes aegypti(71%)和赤拟谷盗Tribolium castaneum(70%)。对该基因在甜菜夜蛾各个发育时期以及不同组织表达量的荧光定量PCR分析表明,SexiCAT基因在甜菜夜蛾各个发育阶段的表达水平存在显著差异,其中成虫期的表达量最高,是卵期表达量的7倍,幼虫期次之,卵期最低;SexiCAT基因在5龄幼虫体壁、中肠、脂肪体和马氏管组织中都有表达,但在脂肪体中表达量最高。甜菜夜蛾SexiCAT基因的成功克隆及同源建模将为今后对其功能研究以及作为靶标设计新型氧化酶抑制剂提供了基础。
Catalase(CAT),one of most important enzymes in organism,plays an essential role in active oxygen metabolism and preventing the formation of free hydroxyl radicals by clearing hydrogen peroxide,superoxide radical and peroxides.In this study,the full-length cDNA of CAT gene from the beet armyworm,Spodoptera exigua,was cloned and characterized by RT-PCR and RACE technique,which is 1 755 bp in length and named as SexiCAT(GenBank accession no.JN051294).The open reading frame(ORF) of SexiCAT is 1 524 bp encoding 507 amino acid residues.The amino acid sequence of SexiCAT shares a significant identity with catelases of Bombyx mori(87%),Drosophila melanogaster(73%),Aedes aegypti(71%),and Tribolium castaneum(70%).The temporal expression profiling revealed that SexiCAT was expressed in different developmental stages of S.exigua at different expression levels,with the highest level in adults,the second highest level in larvae and the lowest level in eggs,and the expression level in adults was 7 times as high as that in eggs.Tissue expression profiling further indicated that in 5th instar larvae of of S.exigua SexiCAT transcript was observed in the integument,midgut,fat body and Malpighian tubules,with the highest expression level in fat body.This study provides a foundation for understanding the functions and applications of CAT as a new target for oxidase blocking agent.