基于转录组金银花WRKY转录因子的挖掘与分析

Mining and Analysis of WRKY Transcription Factors in Transcriptome-based Lonicera japonica

WRKY转录因子是植物抗逆应答过程中最为常见的蛋白家族。为了挖掘金银花抗寒相关的WRKY基因,本研究基于已有的转录组数据,利用植物转录因子数据库和BioEdit对WRKY进行筛选及保守结构域分析,对筛选出的WRKY转录因子利用ExPASy进行理化性质分析,利用PSORT进行亚细胞定位预测,NetPhos进行磷酸化位点分析并且利用MEME数据库对WRKY基因的氨基酸进行序列分析,运用MEGA软件对筛选的金银花WRKY转录因子基因和21个已知抗寒的WRKY基因进行序列同源比对,并构建系统进化树。结果表明:金银花中筛选的30个WRKY基因的蛋白为85～721个氨基酸,理论等电点的范围为4.85到9.76。所有LjWRKY基因编码的蛋白中只有LjWRKY6为稳定蛋白,其余均属于不稳定蛋白,所有蛋白均属于亲水性蛋白。亚细胞定位预测结果,LjWRKY所有蛋白定位于细胞核。除LjWRKY23不具有Try磷酸化位点,其它所有氨基酸均具有Ser、Thr和Try磷酸化位点。LjWRKY蛋白比较保守,WRKYGQK没有发生任何变异,且包含三类WRKY转录因子。系统进化树发现,有12个WRKY基因与已知抗寒基因有同源性,推测其参与了金银花低温逆境下的反应。研究结果可以通过筛选的WRKY基因,进行进一步的转基因验证,进而为植物抗寒分子育种提供分子基础。

WRKY transcrip tion factor is the most common protein family in the process of stress response in plants. In order to explore the WRKY genes of Lonicera japonica related to cold resistance, based on the existing transcriptome data, WRKY was screened using the plant transcription factor database and BioEdit, and the conserved domain analysis was performed. Physicochemical properties of the screened WRKY transcription factors were analyzed using ExPASy. Subcellular localization was predicted by PSORT, NetPhos was used to analyze phosphorylation site analysis and the MEME database was used for sequence analysis of WRKY gene amino acids, the screened Lonicera japonica WRKY transcription factor genes and 21 known cold-resistant WRKY genes were homologous compared by MEGA software, and a phylogenetic tree was constructed. The results showed that the 30 WRKY genes in the Lonicera japonica were 85~721 amino acids and the theoretical isoelectric point ranged from 4.85 to 9.76. Of all the proteins encoded by LJ WRKY gene, only LjWRKY6 was a stable protein, and the rest were all unstable proteins, and all proteins belonged to hydrophilic protein. The subcellular localization predicted that all LjWRKY proteins were located in the nucleus. Except that LjWRKY23 did not have Try Ph acidification sites, all other amino acids had Ser, Thr and Try phosphorylation sites. LjWRKY protein was relatively conserved, WRKYGQK did not have any mutation, and contained three types of WRKY transcription factors. Phylogenetic trees revealed that 12 WRKY genes shared homology with known cold-resistant genes, and it was speculated that they participated in the reaction of Lonicera japonica under low temperature stress. The results of the study could be further verified by screening WRKY genes, which would provide a molecular basis for plant cold-resistant molecular breeding.