铁皮石斛部分WRKY转录因子的基因克隆及研究

发布时间：2018-06-08 21:47

本文选题：WRKY + DoWRKY6　；参考：《西南交通大学》2017年硕士论文

【摘要】：WRKY是植物转录因子家族中较为重要的一员,在植物的生长发育过程中起着极为重要的转录调控作用。本论文基于铁皮石斛原球茎转录组数据,获得了 20个DoWRKY转录因子家族成员的保守区片段,并进一步研究了DoWRKY 基因在不同龄级、茎不同部位及实验室组培幼苗的铁皮石斛材料中的表达情况。通过对20个DoWRKY基因的保守区序列进行系统发育与序列保守性分析,发现这些成员都,有至少一个WRKYGQK构成的氨基酸残基保守结构域,并且通过系统发育树的构建,发现这20个基因亲缘性有较大差异。通过生物信息学分析获得了铁皮石斛DoWRKY6基因的蛋白序列特征,铁皮石斛DoWRKY6基因具有长度为1,017bp的完整ORF,编码339个氨基酸,该蛋白只有一个WRKY结构域和一个结构为CX5C23HX1H的锌指结构,属于II类WRKY转录因子,且WRKY 结构域与锌指结构会和DNA的W盒进行特异性结合。通过Protparam在线预测分析DoWRKY6蛋白的分子式为C1641H2556N454O530S13,相对分子质量为37,541.8,理论等电点为 4.86,总平均亲水性(Grand average of hydropathicity,GRAVY)为-0.756,判定DoWRKY6灰转录因子为疏水性蛋白。利用荧光实时定量PCR技术获得了DoWRKY6基因在不同龄级及茎不同部位的铁皮石斛中的表达特点和在组培幼苗的不同器官中的相对表达量,发现DoWRKY6基因主要在茎中表达,且随着生长年限增长其表达量会大大降低;而在茎中又主要是在茎的芽端部分表达,其次是在茎的基部表达,从而表明DoWRKY6对铁皮石斛茎的生长发育有重要的调控作用。
[Abstract]:WRKY is an important member of the family of plant transcription factors, which plays an important role in the process of plant growth and development. Based on the transcriptome data of protocorm of Dendrobium candidum, the conservative regions of 20 members of DoWRKY transcription factor family were obtained, and the DoWRKY gene was further studied at different age levels. Expression of Dendrobium candidum in different parts of stem and tissue culture seedlings in laboratory. Based on the phylogenetic and conserved analysis of 20 conserved regions of DoWRKY gene, it was found that all of these members had at least one amino acid residues conserved domain of WRKYGQK, and the phylogenetic tree was constructed. It was found that there were significant differences in the genetic relationship of the 20 genes. The protein sequence of Dendrobium candidum DoWRKY6 gene was obtained by bioinformatics analysis. Dendrobium candidum DoWRKY6 gene has a complete ORFwhose length is 1017bp and encodes 339 amino acids. The protein has only one WRKY domain and a zinc finger structure with CX5C23HX1H structure. It belongs to class II WRKY transcription factors, and the WRKY domain binds specifically to the zinc finger structure and the W box of DNA. The molecular formula of DoWRKY6 protein was C1641H2556N454O530S13, the relative molecular weight was 37541.8, the theoretical isoelectric point was 4.86, and the total average hydrophobic average of hydropathicity GRAVYwas -0.756. The grey transcription factor of DoWRKY6 was determined as hydrophobic protein. The expression characteristics of DoWRKY6 gene in Dendrobium candidum at different age levels and different parts of stem were obtained by real-time quantitative PCR. The results showed that DoWRKY6 gene was mainly expressed in stems of Dendrobium candidum in different organs of tissue culture seedlings. The expression of DoWRKY6 was mainly expressed at the end of shoot and then at the base of stem, which indicated that DoWRKY6 played an important role in the growth and development of Dendrobium candidum stem.
【学位授予单位】：西南交通大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：Q943.2;S567.239

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