广藿香香叶醇合酶基因克隆及表达分析

摘要

Geraniol synthase (GES),which is very important to the biosynthesis of geraniol,is one of the key enzymes in terpenoids pathways.Based on the transcriptome analysis of P.cablin,one unique sequence encoding GES was discovered and the primers for PCR were designed from it,the full-length GES cDNA was cloned using RT-PCR strategy and its bioinformatics analysis has been done.Gene expression profile of PcGES1 in leaves and stems of different development stages of four cultivars was evaluated using quantitative reverse-transcription polymerase chain reaction (qRT-PCR).The results showed that the gene GES in P.cablin,named as PcGES1,accession number KF926075 in GenBank,contains a 1 734bp open reading frame and encodes a predicted protein of 577 amino acids.Phylogenetic analysis indicated that the amino acids encoded by GES1 of P.cablin were closest to that of Ocimum basilicum.PcGES protein was located in chloroplast,no transmembrane district,qRT-PCR analysis indicated that PcGES1 mainly expressed in leaves,and the highest expression was in old leaves.The expression pattern analysis of PcGES1 among four cultivars indicated that the expression pattern of P.cablin (Blanco) Benth.cv.shipaiensis was similar to P.cablin (Blanco) Benth.cv.gaoyaoensis,while that of P.cablin (Blanco) Benth.cv.hainangensis was similar to P.cablin (Blanco) Benth.This study provided a foundation for exploring the mechanism of terpenoid biosynthesis in P.cablin plants.%香叶醇合酶(geraniol synthase,GES)是香叶醇形成过程中非常重要的酶,是萜类代谢途径的限速酶.根据课题组广藿香转录组数据中的GES转录本序列设计基因全长扩增引物,采用RT-PCR方法克隆了广藿香GES基因的全长cDNA序列.对该基因进行了相关的生物信息学分析,并利用荧光实时定量PCR法检测了PcGES1基因在4个广藿香栽培种中不同时期茎、叶中的表达情况.结果显示:广藿香GES基因包含一个完整的ORF框,长1 734 bp,编码577个氨基酸,命名为PcGES1,GenBank登录号为KF926075;PcGES1基因编码的氨基酸序列与罗勒GES基因编码的氨基酸序列最为相近.广藿香GES蛋白定位在叶绿体中,无跨膜区域.PcGES1主要在叶中表达,老叶中表达量最高;从不同栽培种来看,PcGES1在石牌广藿香和高要广藿香中表达模式相似,在海南广藿香与印尼广藿香中表达相似,在海南广藿香老叶中表达最高.该研究结果为进一步阐明广藿香萜类代谢途径奠定了基础.