茄子SmWRKY1转录因子基因克隆及序列分析

Cloning and Sequence Analysis of SmWRKY1 Transcription Factor Gene in Eggplant

克隆茄子转录因子SmWRKY1基因,对其序列进行分析,为SmWRKY1基因在茄子抗逆分子育种的应用中提供理论依据。以茄子幼苗为试材,依据GenBank上已提交的番茄、甜椒、烟草的WRKY基因序列设计引物,提取总RNA利用RT-PCR方法扩增得到基因片段,克隆、测序并通过NCBI在线和DNAMAN等软件对其核酸及氨基酸序列进行分析。同源克隆得到SmWRKY1,核酸序列930 bp,翻译为310个氨基酸组成的蛋白序列,其中Ser和Thr量最多,分别占总氨基酸数的12.0%和11.0%。NCBI检索SmWRKY1属于WRKY基因超家族,第114至120个氨基酸含有WRKY保守结构域"WRKYGQK",其锌指结构为C2H2型,初步证实SmWRKY1属于第二大类WRKY转录因子。DNAMAN构建茄子SmWRKY1与不同植物WRKY蛋白的系统发育树,表明茄子WRKY1与番茄中的WRKY1蛋白亲缘关系最近,同源性达96%。WRKY基因在植物耐逆性的研究中有着重要的作用,通过克隆出SmWRKY1基因片段,为进一步克隆其全长、研究其功能做前期准备,为分子育种、培育茄子耐逆性新品种奠定了一定的基础。

The aims were to clone the transcription factor SmWRKY1 gene in eggplant and analyze its sequence characteristics, and provide a theoretical basis for the application of molecular breeding of Sm WRKY1. Eggplant seedlings were used as experimental materials. Specific primers were designed based on the sequences of WRKY gene from tomato, sweet pepper and tobacco, which has been submitted to GenBank. SmWRKY1 gene fragment was amplified by RT-PCR. NCBI online and DNAMAN software were used to analyze nucleic acids and amino acid sequences. SmWRKY1 were cloned through homology-based cloning. SmWRKY1 included 930 nucleotide and encoding 310 amino acids, where Ser and Thr are largest amount of the total number of amino acids with 12.0% and 11.0%. NCBI BLAST showed that SmWRKY1 belonged to WRffY gene superfamily with 114th to 120th amino acids containing WRKY conserved domain ‘WRKYGQK＇ and the zinc finger structure of type C2H2. SmWRKY1 was confirmed belonging to the second category of WRKY superfamily. Plant phylogenetic tree including SmWRKY1 proteins and other plant proteins was built through DNAMAN software, showed that SmWRKY 1 had the closest relationship with tomato WRKY 1 protein with homology of 96%. WRKY gene played an important role in plant stress tolerance and this research cloning gene SmWRKY1 fragment made early preparations for further cloning of its full-length and the studing of itsfunctions and laid a certain foundation for molecular breeding and cultivation of new varieties of eggplant.