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中国农学通报

中国农学通报 ›› 2010, Vol. 26 ›› Issue (12): 5-12.

所属专题： 生物技术； 玉米

• 生物技术科学 • 上一篇    下一篇

玉米螟细胞色素P450家族CYP6AE25的克隆及序列分析

袁辉1,张雨良2,杨峰山3,张一彤3   

  • 收稿日期:2010-01-18 修回日期:2010-02-04 出版日期:2010-06-20 发布日期:2010-06-20
  • 基金资助:

    中央级公益性科研院所基本科研业务费专项

Cloning and Sequence Analysis of Cytochrome P450 CYP6AE25 Gene from Ostrinia furnacalis

Yuan Hui1, Zhang Yuliang2, Yang Fengshan3, Zhang Yitong3   

  • Received:2010-01-18 Revised:2010-02-04 Online:2010-06-20 Published:2010-06-20

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摘要：

摘要：昆虫细胞色素P450是一类在生物代谢中起重要作用的基因家族，承担着许多重要的生理功能，包括对植物次生物质代谢和杀虫剂的解毒等。本研究以玉米螟5龄幼虫总RNA为模板，根据不同昆虫P450基因家族保守氨基酸序列，设计并合成简并引物，利用RT-PCR扩增出了玉米螟细胞色素P450基因的中间片段，通过RACE技术获得玉米螟P450基因家族一员全长序列，将其克隆到pMD19-T载体进行序列测定。测序获得了编码523个氨基酸残基的2315 bp的全长cDNA，命名为OfP450(GenBank登录号：EU807990)。国际P450命名委员会将其定为CYP6AE25。本文还从生物信息学角度对CYP6AE25进行分析，分别从蛋白的理化性质、结构组成、信号肽、跨膜结构域、二级结构、三级结构与功能结构域等作了较为详细的分析与预测。进一步研究其高级结构与功能的关系提供理论依据。CYP6AE25蛋白与其它P450分子进化树分析结果显示，与昆虫CYP6家族同源性较高，与CYP其他家族的遗传距离较远，同源性较低。

关键词: 枯草芽孢杆菌, 枯草芽孢杆菌, 稻曲病菌, 拮抗

Abstract:

Abstract:Insect cytochrome P450 is a kind of gene family that plays important roles in biological metabolism and performs many important physiological functions, including the plant secondary metabolism and insect resistance to insecticides. In this study, total RNA sample of five instar Ostrinia furnacalis larvae midgud was used as template, a pair of degenerate primers were designed based on the reported sequences of conservative amino acid regions of P450 protein family in insects. The cDNA encoding the cytochrome P450 protein was isolated from the five instar Ostrinia furnacalis larva midgut by reverse transcription polymerase chain reaction (RT-PCR) and RACE technology.The complete cDNA fragment was cloned into vector pMD19-T. Sequnence analysis showed that the full-length of Ostrinia furnacalis P450 (GenBank accession no. EU807990) was 2315 bp, encoding 523 amino acid residues. The sequence of OfP450 is named as CYP6AE25 by the International P450 Nomenclature Committee. The research made a more detailed analysis and forecast for CYP6AE25 from the aspects of physical and chemical properties, composition, signal peptide, transmembrane domain, secondary structure, tertiary structure and functional structure domain.It provides a theoretical basis to study between the relationship structure and function. Molecular evolutionary tree analysis revealed that CYP6AE25 protein and other insect family P450 have higher homology, compared with the other CYP family, farther genetic distance, less homologous.