秦艽5-磷酸脱氧木酮糖还原异构酶基因(GmDXR)的克隆和表达分析

Cloning and expression analysis of 1-deoxy-D-xylulose 5-phosphate reductoisomerase gene from Gentiana macrophylla(GmDXR)

目的从药用植物秦艽Gentiana macrophylla中克隆了萜类成分合成途径的关键酶——5-磷酸脱氧木酮糖还原异构酶(1-deoxy-D-xylulose 5-phosphate reductoisomerase,DXR)基因,分析其序列特征及表达方式。方法克隆了秦艽DXR基因(GmDXR),分析了GmDXR编码蛋白序列的特征,并采用实时定量分析了GmDXR的表达模式。结果秦艽GmDXR基因包含1个完整的长1 428 bp的ORF框,编码475个氨基酸;GmDXR与萝芙木、番茄等植物DXR蛋白具有很高的同源性(≥85%),无跨膜结构域、信号肽等结构,主要定位于叶绿体中。定量PCR结果显示,GmDXR主要在秦艽的叶中进行表达,受到植物激素茉莉酸甲酯(methyl jasmonate,MeJA)的诱导。结论 GmDXR含有DXR蛋白的保守结构特征;在秦艽不同器官中GmDXR基因表达不同,且受到MeJA的诱导。该研究为今后研究秦艽环烯醚萜类化合物的生物合成机制提供了依据。

Objective To clone the gene sequence of the key enzyme in synthesis pathway of terpenoids, 1-deoxy-D-xylulose 5-phosphate reductoisomerase （DXR） from Gentiana macrophylla （GmDXR） and to analyze its characteristic and expression. Methods GmDXR gene was cloned and the sequence characterization was analyzed with bioinformatics methods, then the expression patterns of GmDXR were studied by real-time PCR. Results GmDXR gene contained a completed open reading frame of 1 428 bp encoding 475 amino acids, GmDXR has high homology （〉 85%） with DXR proteins from Rauvolfia verticillata, Lycopersicon esculentum, and other plants. Further analysis showed that GmDXR had non-transmembrane domain structure and signal peptide, which mainly located in the chloroplast. Quantitative PCR results showed that GmDXR mainly expressed in the leaves of G. macrophylla and could be induced by plant hormone such as methyl jasmonate （MeJA）. Conclusion GmDXR contains conserved structures of DXR protein. GmDXR expresses variously in different organs ofG. macrophylla and could be induced by MeJA. It is very helpful for the future research on biosynthetic mechanism of iridoid compounds in G. macrophylla.