腊梅葡萄糖-6-磷酸脱氢酶(G6PDH1)基因的克隆及表达分析  

Cloning and expression analysis of glucose-6-phosphate dehydrogenase 1( G6PDH1) gene from Chimonanthus praecox

作　　者：王晓晖[1] 刘晓[1] 高博闻[1,2] 张钟秀[1] 史社坡[1] 屠鹏飞[1] 

机构地区：[1]北京中医药大学中药现代研究中心,北京100029 [2]内蒙古科技大学包头医学院药学院,内蒙古包头014060

出　　处：《中国中药杂志》2015年第21期4160-4164,共5页China Journal of Chinese Materia Medica

基　　金：北京市自然科学基金青年基金项目(7154216)

摘　　要：葡萄糖-6-磷酸脱氢酶是磷酸戊糖途径中的关键限速酶。研究根据已经报道的G6PDH基因的保守序列引物扩增腊梅花中G6PDH基因的核心序列,利用RACE技术从腊梅中首次克隆得到一个G6PDH基因c DNA全长,命名为G6PDH1,并对G6PDH1蛋白质进行细胞定位预测、跨膜结构分析、三维结构模拟及聚类分析,预测G6PDH1的生物学信息;利用实时荧光定量PCR检测G6PDH1在腊梅不同组织中和冷处理条件下的表达差异分析。该研究中克隆获得的G6PDH1基因的全长为2 011bp,开放阅读框1 551 bp,编码516个氨基酸。组织表达分析的结果显示,G6PDH1基因主要在花和根中表达,在叶和茎中表达量较低。通过冷处理,该基因在12 h时达到最高表达水平。研究通过对腊梅花中G6PDH1基因全长c DNA克隆和表达特异性分析,为以后深入研究腊梅的抗寒机制奠定了基础。Glucose-6-phosphate dehydrogenase is main regulatory enzyme for pentose phosphate pathway. To amplify the core sequence of G6 PDH gene from Chimonanthus praecox,the primers were synthesized,based on the conserved nucleotide sequence of other reported plant G6 PDH genes. The specific primers were designed according to the major fragment. The full length c DNA of the G6PDH1 gene was isolated by the 3＇ and 5＇ rapid amplification of c DNA ends approach. Transcript levels of G6PDH1 isoform was measured by real-time quantitative RT-PCR in different tissues and in responds to cold treatment. The G6PDH1 subcellular localization,transmembrane domain,three-dimensional structure,and phylogenetic analysis were predicted by different software to analysis the bioinformatics of G6PDH1 protein. The G6PDH1 c DNA sequence was 2 011 bp in length and consisted of 1 551 bp Open Reading Frame（ ORF）,encoding a protein of 516 amino acids. Expression analysis results in different tissues showed that G6PDH1 was primarily observed in flowers and roots,as opposed to the leaves and stems. Cold treatment experiments indicated that cold treatment caused a rapid increase in G6PDH1 expression in flowers within 12 h. The full-length c DNA of G6PDH1 and its expression analysis will play an important role for further study on cold stress responses in Ch. praecox.

关 键 词：腊梅 G6PDH基因 表达分析 

分 类 号：S567.19[农业科学—中草药栽培]