维药新塔花(Ziziphora bungena)zbHDS基因克隆及组织表达分析

Cloning and Tissue Expression Analysis of zbHDS in Uighur Medicial Plant(Ziziphora bungena)

对维药新塔花(Ziziphora bungena)萜类化合物生物合成途径中,关键酶1-羟基-2甲基-2-(E)-丁烯基-4-二磷酸合成酶HDS基因克隆,进行组织表达特异性分析,为研究新塔花萜类化合物生物合成途径与基因调控提供基础。结合新塔花转录组数据库,根据编码区序列设计引物,通过RT-PCR方法对Z. bungena基因HDS (zbHDS)的编码区cDNA进行克隆,对其进行相关生物信息学分析,通过Real-Time进一步分析其组织表达特异性。RT-PCR扩增了一个长2 465 bp的zbHDS (登录号:MG065856)基因片段,含一个2 229 bp的开放阅读框,编码742个氨基酸。进化树分析结果显示,与丹参(Salvia miltiorrhiza)具有较近的亲缘关系。Real-Time PCR结果显示zbHDS基因在各器官中均有表达,在茎和叶中表达量较高,根和花中表达量相对较低。本研究首次从新塔花中克隆HDS基因并获得其编码区序列,zbHDS基因具有组织表达特异性,为进一步研究新塔花中萜类化合物生物合成途径提供数据支持。

To provide the basis for the further studies on biosynthesis and gene regulation of torpedoed,we tried to clone 1-hydroxy-2-methyl-2-(E)-butenyl-4-diphosphate synthase(HDS)gene that plays an important role in torpedoed biosynthesis pathway in Uighur Medicial plant(Ziziphora bungena).According to the Zb HDS gene sequences of transcriptome of Z.bungena,a pair of primers were designed and the ORF of c DNA of sequences was obtained by RT-PCR,Then TA cloning,sequencing,and sequence analysis were performed in this research.The expression of Zb HDS in different organ of Z.bungena was detected by Real-Time PCR.The Zb HDS gene was obtained in length of2 465 bp gene fragments(GenBank accession number MG065856),with an open reading frame of 2 229 bp in length,encoding 742 amino acids.Phylogenetic analysis on the amino acid sequence of Zb HDS with those of other plants showed that Zb HDS was closely related to Salvia miltiorrhiza.The result of Real-Time PCR showed that Zb HDS gene was expressed in different tissues of Z.bungena.It showed that Zb HDS was high expression in leaves and stems while low expression in root and flower.The Zb HDS gene is cloned from Z.bungena for the first time and the expression of Zb HDS is tissue-specific.This research lays a foundation for studying the gene expression pattern and regulating the functions of Zb HDS in torpedoed biosynthesis.