小麦光温敏雄性不育系BS366铜锌超氧化物歧化酶基因的克隆及表达分析
发布时间：2018-11-28 11:49
【摘要】：超氧化物歧化酶(SOD,superoxide dismutase)是植物中一种主要的抗氧化酶,在植物应对逆境胁迫及抗衰老中起重要作用。本研究从基因芯片数据中筛选获得小麦Cu/Zn-SOD基因的EST序列,通过序列比对后拼接得到小麦Cu/Zn-SOD的候选基因,利用PCR技术在小麦光温敏雄性不育材料BS366中克隆并获得该基因。通过对Cu/Zn-SOD基因序列进行生物信息学分析,结果表明,该基因拥有连续且完整的开放阅读框,长495bp,编码164个氨基酸。氨基酸序列分析发现,该蛋白具有保守的Cu/Zn-SOD功能结构域与典型的Cu/Zn-SOD三维结构,且定位于细胞质中。通过同源进化分析表明,该蛋白与二穗短柄草(Brachypodium distachyon(L.)Beauv.)和大麦(Hordeum vulgare L.)的Cu/Zn-SOD蛋白亲缘关系较近,相似度分别为89%和94%。利用实时荧光定量PCR技术对其在小麦不同组织的表达特异性及不同逆境胁迫下的表达模式进行分析,结果表明,该基因在根、茎、叶、雌蕊、雄蕊、颖壳中均有表达,属于组成型表达,且在小麦的地上部含叶绿体的组织中含量较高;同时受多种胁迫诱导,可能参与了多种胁迫诱导调控途径。通过对该基因在不同育性环境中BS366育性转换期花药中的表达模式分析,发现可育环境下,在小孢子母细胞时期和减数分裂期的表达量分别约为对照的8倍与16倍;而不育环境下,该基因表达水平无明显变化。因而推测,小麦Cu/Zn-SOD基因可能参与了光温敏雄性不育系BS366的育性调控。本研究为深入研究Cu/Zn-SOD基因在小麦中的作用机理奠定了重要基础。
[Abstract]:Superoxide dismutase (SOD,superoxide dismutase) is a major antioxidant enzyme in plants, which plays an important role in coping with stress and anti-senescence. In this study, the EST sequence of wheat Cu/Zn-SOD gene was screened from gene chip data, and the candidate gene of wheat Cu/Zn-SOD was obtained by sequence alignment. The PCR technique was used to clone and obtain the gene from wheat photo-thermosensitive male sterile line BS366. The results of bioinformatics analysis of Cu/Zn-SOD gene showed that the gene had a continuous and complete open reading frame with a length of 495bpand encoding 164 amino acids. Amino acid sequence analysis showed that the protein had a conserved Cu/Zn-SOD functional domain and a typical Cu/Zn-SOD three-dimensional structure, and was located in the cytoplasm. The result of homologous evolution analysis showed that the protein was similar to (Brachypodium distachyon (L.) Beauv.). Barley (Hordeum vulgare L.) The similarity of Cu/Zn-SOD protein was 89% and 94%, respectively. The expression specificity of the gene in different wheat tissues and its expression patterns under different stresses were analyzed by real-time fluorescence quantitative PCR. The results showed that the gene was expressed in roots, stems, leaves, pistil, stamens and glumes. It belongs to the constitutive expression, and the content of chloroplast is higher in the tissue with chloroplast in the aboveground part of wheat. At the same time, it may be involved in the regulation of multiple stress induced by multiple stresses. By analyzing the expression pattern of the gene in the anthers of BS366 fertility transition in different fertility environments, it was found that in fertile environment, the expression of the gene in microspore mother cell stage and meiosis stage was about 8 times and 16 times as much as that of the control, respectively. However, there was no significant change in the expression level of the gene in infertile environment. It is suggested that the Cu/Zn-SOD gene may be involved in the fertility regulation of the photo-temperature sensitive male sterile line BS366. This study laid an important foundation for further study on the mechanism of Cu/Zn-SOD gene in wheat.
【作者单位】： 北京农学院植物科学技术学院;北京市农林科学院杂交小麦工程技术研究中心;杂交小麦分子遗传北京市重点实验室;首都师范大学生命科学学院;
【基金】：国家科技支撑计划项目(2013BAD04B01,2014BAD01B09) 国家自然科学基金(31371699) 北京市自然科学基金项目(5131001) 北京市农林科学院科技创新能力建设专项(KJCX20140405);北京市农林科学院青年基金(QNJJ201427) 北京市科技计划项目(Z141100002314018)
【分类号】：S512.1
本文编号：2362745
[Abstract]:Superoxide dismutase (SOD,superoxide dismutase) is a major antioxidant enzyme in plants, which plays an important role in coping with stress and anti-senescence. In this study, the EST sequence of wheat Cu/Zn-SOD gene was screened from gene chip data, and the candidate gene of wheat Cu/Zn-SOD was obtained by sequence alignment. The PCR technique was used to clone and obtain the gene from wheat photo-thermosensitive male sterile line BS366. The results of bioinformatics analysis of Cu/Zn-SOD gene showed that the gene had a continuous and complete open reading frame with a length of 495bpand encoding 164 amino acids. Amino acid sequence analysis showed that the protein had a conserved Cu/Zn-SOD functional domain and a typical Cu/Zn-SOD three-dimensional structure, and was located in the cytoplasm. The result of homologous evolution analysis showed that the protein was similar to (Brachypodium distachyon (L.) Beauv.). Barley (Hordeum vulgare L.) The similarity of Cu/Zn-SOD protein was 89% and 94%, respectively. The expression specificity of the gene in different wheat tissues and its expression patterns under different stresses were analyzed by real-time fluorescence quantitative PCR. The results showed that the gene was expressed in roots, stems, leaves, pistil, stamens and glumes. It belongs to the constitutive expression, and the content of chloroplast is higher in the tissue with chloroplast in the aboveground part of wheat. At the same time, it may be involved in the regulation of multiple stress induced by multiple stresses. By analyzing the expression pattern of the gene in the anthers of BS366 fertility transition in different fertility environments, it was found that in fertile environment, the expression of the gene in microspore mother cell stage and meiosis stage was about 8 times and 16 times as much as that of the control, respectively. However, there was no significant change in the expression level of the gene in infertile environment. It is suggested that the Cu/Zn-SOD gene may be involved in the fertility regulation of the photo-temperature sensitive male sterile line BS366. This study laid an important foundation for further study on the mechanism of Cu/Zn-SOD gene in wheat.
【作者单位】： 北京农学院植物科学技术学院;北京市农林科学院杂交小麦工程技术研究中心;杂交小麦分子遗传北京市重点实验室;首都师范大学生命科学学院;
【基金】：国家科技支撑计划项目(2013BAD04B01,2014BAD01B09) 国家自然科学基金(31371699) 北京市自然科学基金项目(5131001) 北京市农林科学院科技创新能力建设专项(KJCX20140405);北京市农林科学院青年基金(QNJJ201427) 北京市科技计划项目(Z141100002314018)
【分类号】：S512.1
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