铁皮石斛多聚泛素基因Polyubiquitin1(DoUb1)的克隆及表达分析  

Cloning and expression analysis of a Polyubiquitin gene (DoUb1) in the Dendrobium officinale Kimura et Migo

作　　者：裴薇[1] 梁易[1] 范静[1] 安红强[1] 王万军[1] 

机构地区：[1]西南交通大学生命科学与工程学院,成都610031

出　　处：《生物学杂志》2017年第3期6-10,共5页Journal of Biology

基　　金：国家自然科学基金项目(31371232)

摘　　要：采用RACE(rapid amplification of cDNA ends)技术获得铁皮石斛(Dendrobium officinale Kimura et Migo)Polyubiquitin1(DoUb1),该基因全长1985 bp,开放阅读框为1371 bp,编码457个氨基酸残基的亲水性蛋白,分子质量为51 179.7 ku,理论等电点p I为7.76;亚细胞定位分析表明该基因产物主要在细胞质或线粒体基质中发挥作用。DoUb1与水稻、玉米及谷子等单子叶植物的多聚泛素基因亲缘关系较近。实时荧光定量PCR(RT-qPCR)分析结果显示,DoUb1在铁皮石斛幼苗根和叶中表达量较高,茎中表达量较低;在干旱胁迫和温度胁迫过程中的表达量都低于正常状况,在干旱胁迫的早期,表达量降低,6 h时达到最低,之后随胁迫时间的延长而缓慢升高;在温度胁迫情况下,DoUb1具有与干旱胁迫相近的响应特性,但低温胁迫相比高温胁迫下,DoUb1表达量更低,表明该基因对低温胁迫更敏感,结果为后期研究铁皮石斛生长发育提供一定基础。Polyubiquitin! （DoUbl） in Dendrobium officinale Kimura et Migo was achieved by RACE （rapid amplification of cDNA ends）. DoUbl contained 1985 bp nucleotides with a putative open reading frame encoding 457 amino acid residues. The predicted molecular weight and theoretical isoelectric point of pI were 51 179. 7 ku and 7. 76 respectively. Subcellular localization analysis showed that gene product played a major role in the cytoplasm or the mitochondrial matrix. Phylogenetic analysis showed that the DoUbl has the closest relativon with monocotyledonous plants such as Zea mays, Setaria italica and Oryza sativa. Results of real-time quantitative PCR （RT-qPCR） showed that the expression of DoUbl was higher in roots and leaves than in stems; DoUbl was down-reg- ulated during drought stress and temperature stress, the expression level was reduced in the early stage of stresses and then slowly in- creased. For drought stress, and the expression level was at the lowest after PEG treatment for 6 h; for temperature stress, it had simi- lar responsive characteristics to drought stress, but in the low temperature stress, the expression of DoUbl was lower than in high tem- perature stress. These results showed that the gene was more sensitive to low temperature stress.

关 键 词：RACE 铁皮石斛 POLYUBIQUITIN RT-QPCR 

分 类 号：Q785[生物学—分子生物学] Q786