香樟IDI基因克隆及表达分析  

Cloning and Expression Analysis of IDI Gene from Cinnamomum camphora

作　　者：刘伟 曹先爽 王进 张瑶瑶 王煜炜 Liu Wei;Cao Xianshuang;Wang Jin;Zhang Yaoyao;Wang Yuwei(SFA Key Laboratory of Bamboo and Rattan Science and Technology,International Centre for Bamboo and Rattan,Beijing,100102)

机构地区：[1]国际竹藤中心,国家林业局竹藤科学与技术重点实验室,北京100102

出　　处：《分子植物育种》2020年第8期2520-2526,共7页Molecular Plant Breeding

基　　金：国家林业局948项目(2014-4-33);“十二五”国家科技支撑计划项目(2012BAD23B03)共同资助。

摘　　要：为了克隆香樟异戊烯基焦磷酸异构酶(isopentenyl diphosphate isomerase,IDI)基因,并进行生物信息学分析,为香樟植物萜类化合物的生物合成研究提供基础。本研究以香樟植物为试样,在转录组测序数据的基础上设计引物,应用RT-PCR技术克隆香樟IDI基因,通过实时定量PCR方法检测IDI基因在香樟根、茎和叶中表达量,利用生物信息学方法对IDI基因编码的蛋白进行表征。结果表明,香樟IDI基因长度868 bp,开放读码框(ORF)为708 bp,编码235个氨基酸,编码蛋白的分子量为27.04 kD,等电点为5.13,蛋白的二级序列结构主要为α-螺旋。经预测香樟IDI是一个定位于细胞质,不含信号肽的亲水性稳定蛋白,具有异戊烯基焦磷酸异构酶的特征结构域。系统进化树分析表明,香樟IDI与野生型马来西亚蕉的亲缘关系较近。实时定量PCR结果表明,IDI在香樟茎中的表达量高于根和叶。本研究成功从香樟中克隆了IDI基因并进行了表征分析,为深入研究香樟IDI基因在萜类合成中的功能提供帮助。To clone the isoamyl pyrophosphate isomer(IDI)gene of Cinnamomum camphora and analyze it by bioinformatics,so as to provide a basis for the biosynthesis of terpenoids from C.camphora.Using C.camphora as sample in this study,primers were designed on the basis of sequencing data of transcriptional group.IDI gene of C.camphora was cloned by RT-PCR,and the expression of IDI gene in root,stem and leaf of C.camphora was detected by real-time quantitative PCR.Then,the protein encoded by IDI gene was analyzed by bioinformatics method.The results showed that the length of C.camphora IDI gene was 868 bp,open reading frame(ORF)encoding 235 amino acids,and the molecular weight of the encoded protein was 27.04 kD,isoelectric point of 5.13.Theα-helix was the main secondary structure in CcIDI.CcIDI was predicted as a hydrophilic and stable protein without signal peptide,which located in the cytoplasm.CcIDI contained a characteristic structural domain of isoprenyl diphosphate isomerase.Phylogenetic tree analysis showed that CcIDI was closely related to Musa acuminata subsp.Malaccensis and Phoenix dactylifera.The results of real-time quantitative PCR showed that the expression of IDI in C.camphora stems was higher than that in roots and leaves.In brief,CcIDI gene was successfully cloned from C.camphora and the CcIDI protein was characterized by bioinformatics method.This study is helpful to further research the function of CcIDI gene in terpenoids synthesis of C.camphora.

关 键 词：香樟(Cinnamomum camphora (L.) Presl) 异戊烯基焦磷酸异构酶 基因克隆 生物信息学分析 

分 类 号：S792.23[农业科学—林木遗传育种]