家蚕细胞色素P450基因CYP9A22的克隆及在中肠和脂肪体的诱导转录  

Cloning of the CYP 9A22 of Cytochrome P450 Genes From the Silkworm,Bombyx mori and Induced Transcription of CYP 9A22 in Fat Body and Midgut

作　　者：赵华强[1] 王东[1] 李兵[1,2] 许雅香[1,2] 沈卫德[1,2] 

机构地区：[1]苏州大学基础医学与生物科学学院,江苏苏州215123 [2]苏州大学蚕桑研究所,江苏苏州215123

出　　处：《蚕业科学》2008年第4期634-641,共8页ACTA SERICOLOGICA SINICA

基　　金：国家重点基础研究发展计划"973"项目(编号2005CB12-1005);国家高技术研究发展计划"863"项目(编号2006-AA10A118)

摘　　要：细胞色素P450对昆虫的生长发育、环境适应性以及抗药性具有重要作用。为了探讨氯氰菊酯对家蚕(Bombyxmori)细胞色素P450基因转录的诱导作用,采用RT-PCR技术和cDNA末端快速扩增(RACE)策略,从家蚕5龄幼虫克隆到细胞色素P450基因CYP9A22(GenBank登录号:EF535804)。CYP9A22基因的cDNA编码区长1 596 bp,编码531个氨基酸,推定的蛋白质分子质量为61 kD,等电点为7.71。将该基因的cDNA和家蚕基因组序列比对,有9个内含子;与已知的CYP9家族的棉铃虫(Helicoverpa armigera)CYP9A14基因的相应氨基酸同源性达到62.3%。用半定量RT-PCR方法分析氯氰菊酯诱导家蚕CYP9A22表达水平,结果表明:CYP9A22的mRNA转录表达具有组织特异性,在中肠的表达量高于脂肪体;氯氰菊酯诱导家蚕24 h,在其脂肪体的表达量是未诱导家蚕脂肪体的3.1倍,初步推测CYP9A22的过量表达可能与抗药性有一定关系。Cytochrome P450 has been proved to be involved in growth and development, environment adaptation and insecticide resistance in insect. In order to illustrate the inducement effect of cypermethrin treatment on the transcription of cytochrome P450 genes in Bombyx mori, CYP9A22 （GenBank accession number： EF535804） from 5th instar larvae was cloned by methods of RT-PCR and RACE. The coding region of CYFgA22 cDNA is 1 596 bp, which encodes 531 amino acid residues with the predicted molecular weight of 61 kD and isoelectric point of 7.71. Alignment result of the cDNA with the silkworm genomic sequences revealed that there are 9 introns in this gene. The deduced amino acid sequence showed 62.3% identity to CYP9A14 from Helicoverpa armigera. The inducement effect of cypermethrin treatment on the transcription of CYFgA22 were determined by semi-quantitative RT-PCR method. The results showed that the transcription level was higher in fat body than in midgut of the 5th instar larvae. The induced transcription level in fat body by the cypermethrin treatment for 24 h was 3.1 times of the control, suggested that overtranscription level of CYFgA22 maybe involved in pyrethroid resistance of Bombyx mori.

关 键 词：家蚕 细胞色素P450 CDNA末端快速扩增 基因克隆 半定量RT-PCR 

分 类 号：S881.2[农业科学—特种经济动物饲养] Q78[农业科学—畜牧兽医]