刊期:双月刊
主管单位:中国科学院
主办单位:中国科学院动物研究所,中国昆虫学会
地址:北京市朝阳区北辰西路1号院5号中国科学院动物研究所
邮编:100101
电话:010-64807137
传真:010-64807137
E-Mail:entom@ioz.ac.cn
刊号:ISSN 2095-1353
        CN 11-6020/Q
国内发行代号:2-151
国际发行代号:BM-407
发行范围:国内外公开发布
定价:138元/册
定价:828元/年
银行汇款:中国工商银行北京海淀西区支行
户名:中国科学院动物研究所
帐号:0200 0045 0908 8125 063

您所在位置:首页->过刊浏览->2020年57卷第4期



苹果蠹蛾NAPDH-细胞色素P450还原酶 (CPR)基因的克隆及表达模式分析
Molecular cloning and expression profiles of NADPH-cytochrome P450 reductase (CPR) gene in the codling moth, Cydia pomonella
李沛蓉;陈高满;杨雪清
点击:246次 下载:14次
DOI:10.7679/j.issn.2095-1353.2020.087
作者单位:沈阳农业大学植物保护学院; 辽宁省经济与应用昆虫教育厅重点实验室
中文关键词:苹果蠹蛾;细胞色素P450还原酶;解毒代谢;抗药性;基因克隆
英文关键词:Cydia pomonella; NADPH cytochrome P450 reductase; detoxification; insecticide resistance; gene cloning
中文摘要:
【目的】 本研究通过克隆苹果蠹蛾Cydia pomonella CPR (CpCPR) 基因cDNA序列,并对该基因的表达模式进行分析,为深入研究P450酶系在苹果蠹蛾对植物次生物质和杀虫剂解毒代谢过程中的作用提供理论基础。【方法】 以近缘昆虫的CPR氨基酸序列作为询问序列,在苹果蠹蛾转录组(SRX371333)中进行筛查比对,获得了苹果蠹蛾CPRCpCPR)基因的cDNA序列。采用RT-PCR技术克隆目的基因的开放阅读框(ORF)。利用生物信息学软件分析目的基因的序列特征、3D结构和与其他昆虫CPR基因的系统进化关系。采用RT-qPCR技术测定CpCPR基因在苹果蠹蛾不同发育阶段(卵、幼虫、蛹和成虫)及幼虫不同组织部位(头部、表皮、脂肪体、中肠和马氏管)的表达水平。【结果】 克隆获得的苹果蠹蛾CpCPR基因的ORF为2 052 bp,编码683个氨基酸残基,预测的蛋白质分子量(Mw)为77.326 ku,理论等电点(pI)为5.65。CpCPR包含FMN区域、NADPH区域和FAD等昆虫CPR的典型特征。系统发育分析表明,苹果蠹蛾CpCPR与鳞翅目昆虫CPR基因聚在一枝。RT-qPCR结果表明,CpCPR基因在苹果蠹蛾的整个发育阶段均有表达,在幼虫期表达量最高;CpCPR基因在4龄幼虫的各部位均有表达,在中肠中的表达量最高。【结论】 克隆获得了苹果蠹蛾CpCPR基因的ORF序列,该基因在苹果蠹蛾主要取食阶段和消化器官中高表达,表明其可能在苹果蠹蛾对植物次生物质和杀虫剂解毒代谢过程中扮演重要作用。
英文摘要:
[Objectives]  In order to improve understanding of the role of cytochrome P450 in the detoxification of plant secondary substances and insecticides, the CPR gene (CpCPR) of the codling moth, Cydia pomonella, was cloned and its expression profile was analyzed. [Methods]  The amino acid sequences of CPR genes from closely-related insect species were used as queries to search against the C. pomonella transcriptome database (SRX371333) and obtain the sequence of the C. pomonella CPR gene. RT-PCR was then used to amplify the Open Reading Frame (ORF) of the target gene. Bioinformatics software was used to compare the sequence characteristics, 3D structure and phylogenetic relationship of CpCPR to those of CPR from other insects. Real-time quantitative PCR (RT-qPCR) was used to determine the relative expression level of CpCPR in different developmental stages (eggs, 1-5 instar larvae, pupae, and adults) and in various tissues (head, cuticle, fat body, midgut, and Malpighian tubes). [Results]  The length of the CpCPR gene is 20 252 bp, which encodes 683 amino acids with a predicted molecular mass of 77.326 ku and a theoretical isoelectric point of 5.65. CpCPR contains typical insect CPR gene characteristics, such as an FMN region, an NADPH region and a FAD region. Phylogenetic analysis indicates that CpCPR clusters with the CPR gene of other Lepidoptera. RT-qPCR results show that the CpCPR gene is expressed in all developmental stages but is most abundant in the larval stage. In addition, CpCPR mRNA was detected in all tissues but was most abundant in the midgut in fourth-instar larvae. [Conclusion]  The ORF of the CpCPR gene was successfully cloned. This gene was most highly expressed during the larval stage and in the midgut of C. pomonella, indicating that it may play an important role in the detoxification of plant secondary substances and insecticides.
读者评论

      读者ID: 密码:   
我要评论:
版权所有©2021应用昆虫学报》编辑部 京ICP备10006425号
本系统由北京菲斯特诺科技有限公司设计开发
您是本站第5520348名访问者