生物学杂志

生物学杂志

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

  

  1. 西南交通大学 生命科学与工程学院， 成都 610031
  • 出版日期:2017-06-18 发布日期:2017-06-18
  • 通讯作者: 王万军，教授，主要从事植物发育生物学研究，E-mail: wanjunwang@home.swjtu.edu.cn
  • 作者简介:裴薇，硕士研究生，主要从事植物发育生物学研究，E-mail: 1141217319@qq.com
  • 基金资助:
    国家自然科学基金项目(31371232)

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

  1. College of Life Science and Engineering, Southwest Jiaotong University, Chengdu 610031, China
  • Online:2017-06-18 Published:2017-06-18

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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pI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。

关键词: RACE, 铁皮石斛, Polyubiquitin, RT-qPCR

Abstract:  Polyubiquitin1 (DoUb1) in Dendrobium officinale Kimura et Migo was achieved by RACE (rapid amplification of cDNA ends). DoUb1 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 DoUb1 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 DoUb1 was higher in roots and leaves than in stems; DoUb1 was down-regulated during drought stress and temperature stress, the expression level was reduced in the early stage of stresses and then slowly increased. For drought stress, and the expression level was at the lowest after PEG treatment for 6 h; for temperature stress, it had similar responsive characteristics to drought stress, but in the low temperature stress, the expression of DoUb1 was lower than in high temperature stress. These results showed that the gene was more sensitive to low temperature stress.

Key words: RACE, Dendrobium officinale Kimura et Migo, Polyubiquitin, RT-qPCR