生物学杂志

生物学杂志 ›› 2020, Vol. 37 ›› Issue (2): 18-.doi: 10.3969/j.issn.2095-1736.2020.02.018

• 研究报告 • 上一篇    下一篇

铁皮石斛FCA基因的表达及生物信息学分析

  

  1. 西南交通大学 生命科学与工程学院， 成都 610031
  • 出版日期:2020-04-18 发布日期:2020-04-17
  • 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:李良建，硕士研究生，主要从事植物发育生物学研究，E-mail:1411673930@qq.com
  • 基金资助:
    国家自然科学基金项目（31371232）

Expression and bioinformatics analysis of FCA gene in Dendrobium officinale Kimura et Migo

  1. School of Life Science and Engineering, Southwest Jiaotong University, Chengdu 610031, China
  • Online:2020-04-18 Published:2020-04-17

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摘要： FCA（FLOWERING CONTROL LOCAL A）基因是自主开花途径中促进植物开花的保守基因。通过电子克隆和末端快速扩增技术 (Rapid amplification of cDNA ends，RACE) 从铁皮石斛中克隆得到FCA基因全长cDNA序列，命名为DoFCA。生物信息学分析显示DoFCA基因开放阅读框为2301 bp，包含19个外显子，18个内含子，编码766个氨基酸残基的蛋白，在N-端（131~210、220~298）有两个RRM结构域，C-端（605~634）存在一个WW结构域。启动子分析发现，该基因启动子区存在与分生组织特异激活、胚乳表达、花药特异基因的表达调控密切相关的调控元件。蛋白序列同源性分析表明，保守区域主要集中在两个RRM结构域和WW结构域部分，表明该蛋白在不同物种中功能保守。系统发育分析显示DoFCA蛋白与石斛类FCA蛋白的进化关系最近，并且发现FCA蛋白在单子叶和双子叶植物进化过程中出现了分化。对DoFCA-WW/FY-PPLPP复合物的分析发现WW结构域中的14Tyr、16Tyr、23Ser及25Trp 4个位点与FY-PPLPP相互作用最紧密，并且23Ser和FY-PPLPP基序的第2个Pro残基形成氢键。逆转录实时荧光定量PCR（RT-qPCR）分析结果显示，DoFCA在铁皮石斛花组织中的表达有优势，在根中表达最少，茎、叶组织中表达量其次。在低温处理12 h时后，其表达量迅速下降，随后表达量又逐渐升高，最后维持在较高水平。表明该基因与铁皮石斛花的发生和形成可能具有重要作用；在低温诱导开花过程中也发挥作用。

关键词: RACE, 铁皮石斛, FCA基因, 实时定量PCR

Abstract: The FCA (FLOWERING CONTROL LOCAL A) gene is a conserved gene that promotes plant flowering in the autonomous pathway. The full-length cDNA sequence of FCA gene was cloned from Dendrobium officinale Kimura et Migo by siloco PCR and rapid amplification of cDNA ends technology (RACE), and named as DoFCA. Bioinformatics analysis revealed that the open reading frame of the DoFCA gene was 2301 bp, containing 19 exons and 18 introns. The DoFCA encoded a protein with 766 amino acid residues which had two RRM domains at N-terminal (131-210, 220-298) and one WW domain at C-terminal (605-634). Promoter analysis revealed that there were regulatory elements in the promoter region of this gene, which were closely related to meristem-specific activation, endosperm expression and expression regulation of anther-specific genes. Protein sequence homology analysis showed that the conserved region mainly concentrated in both the RRM and WW domain, which indicated that the protein was functionally conserved in different species. Phylogenetic analysis showed that DoFCA protein had the closest evolutionary relationship with Dendrobium-like FCA protein, and it was found that FCA protein differentiated during the evolution of monocotyledons and dicotyledons. Analysis of the DoFCA-WW/FY-PPLPP complex revealed that the four sites (14Tyr, 16Tyr, 23Ser, and 25Trp) in the WW domain interacted most closely with FY-PPLPP motif, and the 23Ser residue formed a hydrogen bond with the second Pro of FY-PPLPP motifs. The results of RT-qPCR analysis showed that the expression level of DoFCA was higher in flower, followed by stem and leaf, and the lowest in root. Under low-temperature stress, the expression level decreased rapidly after treated for 12 h, then increased gradually, and finally maintained at a high level. These results suggested that the gene might play an important role in the flower development and formation of Dendrobium officinale Kimura et Migo and also in the process of low temperature induced flowering.

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

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