三七甲羟戊酸激酶PnMVK1基因的克隆和生物信息学分析

Cloning and analysis of mevalonate kinase(PnMVK1) gene in Panax notoginseng

药用植物三七通过甲羟戊酸途径(mevalonic acid pathway)合成三萜皂苷生物合成的前体。甲羟戊酸激酶(mevalonate kinase,MVK)则是甲羟戊酸合成途径中ATP依赖的限速酶之一。本研究根据课题组已获得的三七[Panax notoginseng(Burk.)F.H.Chen]转录组数据中的MVK1转录本序列,利用RT-PCR方法获得三七MVK1(PnMVK1)基因的全长cDNA序列;对PnMVK1蛋白进行理化性质、蛋白二级结构及三维结构预测分析,并预测了该蛋白的结构与功能;利用实时荧光定量PCR方法检测了PnMVK1基因在三七的根、茎、叶、花中的表达情况。序列分析表明,克隆获得的三七PnMVK1基因长为1 164 bp,编码387个氨基酸,GenBank登录号JQ957844。生物信息学预测PnMVK1蛋白不含跨膜区,不含信号肽,具有GHMP激酶等保守结构域。PnMVK1基因在三七的根中表达丰度最高。本研究成功克隆并分析了三七MVK1的全长序列,为进一步阐明三七三萜代谢途径奠定基础。

Mevalonate kinase（MVK） is a key enzyme in plant terpenoid biosynthetic pathway.This study focused on cloning and analysis of Panax notoginseng（Burk.） F.H.Chen MVK（PnMVK1） gene.After searching the transcriptome dataset of P.notoginseng,one unique sequence encoding MVK was discovered.The primers were designed according to the transcript sequence of PnMVK1 from the P.notoginseng transcriptome dataset.And then,the open reading frame of PnMVK1 was cloned from P.notoginseng by using RT-PCR strategy.The physical and chemical properties,secondary structure and three-dimensional structure of the PnMVK1 protein were forecasted and analyzed,and its structure and function were predicted.The cDNA（named as PnMVK1） contains a 1 164 bp open reading frame and encodes a predicted protein of 387 amino acids.The GenBank accession number for this gene is JQ957844.No transmembrane region and signal peptide were present in PnMVK1.The conserved domain of mevalonate kinase was present in PnMVK1.PnMVK1 was more abundant in P.notoginseng root than other organisms.This study cloned and analyzed PnMVK1 gene from P.notoginseng for the first time.The result will provide a foundation for exploring the mechanism of terpenoid biosynthesis in P.notoginseng plants.