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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。
As an important substance in vivo, catalase (CAT) plays an important role in the metabolism of reactive oxygen species and plays an important role in clearing H2O2, superoxide radicals and peroxides and preventing the formation of hydroxyl radicals . In this study, the CAT gene of Spodoptera exigua (Hübner) was cloned and analyzed by RT-PCR and RACE for the first time. The GenBank accession number was JN051294. The full-length cDNA of cNDA was 1755bp and the open reading frame was 1524bp. Presumably encode 507 amino acids. The amino acid sequence alignment showed that the polypeptide sequence was highly conserved. The sequence identity with CAT of other insects was Bombyx mori (87%), Drosophila melanogaster (73%), Aedes aegypti (71) %) And Tribolium castaneum (70%). Quantitative real-time PCR analysis of the expression level of SexiCAT gene at different developmental stages of Beet armyworm showed that the expression level of SexiCAT gene was significantly different at all developmental stages of Beet armyworm. Among them, the expression level of SexiCAT gene was highest in egg stage 7 times of that in larvae, followed by larvae. The expression of SexiCAT gene in the body wall, midgut, fat body and M. tuberculosis tissues of 5th instar larvae was the highest, but highest in fat body. The successful cloning and homology modeling of the SexiCAT gene of beet armyworm will provide the basis for its function research and target design of novel oxidase inhibitors in the future.