大豆转录因子GmWIN1-1的鉴定与表达分析

Identification and Expression Analysis of Soybean Transcription Factor GmWIN1-1

为挖掘参与大豆胁迫应答的调控基因,研究聚焦大豆转录因子WAX INDUCER1(WIN1/SHN1)家族成员的鉴定和功能分析,以拟南芥AtWIN1编码序列为索引,在大豆转录组数据库中进行Blast比对,鉴定获得1条编码大豆GmWIN1-1转录因子的cDNA序列。通过生物信息学方法和qRT-PCR技术分析GmWIN1-1转录因子的序列特征及其在大豆各组织中和干旱胁迫下的表达水平。结果表明,该序列全长570 bp,编码189个氨基酸,蛋白分子质量为21.2 ku,理论等电点为8.45,为亲水性蛋白。GmWIN1-1蛋白二级结构主要元件为无规则卷曲,三级结构建模与模板5wx9.1蛋白氨基酸序列相似性为45%,为单体蛋白。多序列比对揭示GmWIN1-1与AtWIN1相似性最高。保守基序分析表明,WIN1蛋白具有3个基序,即AP2保守基序、中间基序、C端基序。系统发育分析显示,GmWIN1-1与MtWIN1亲缘关系较近。PlantCARE软件预测,GmWIN1-1启动子含有多个响应逆境胁迫的核心顺式元件。实时荧光定量PCR显示,GmWIN1-1在大豆各组织中的表达水平差异显著,在花器官中表达量最高。干旱胁迫条件下,GmWIN1-1的表达水平呈现先小幅度下降而后快速上调再下降的趋势。大豆GmWIN1-1转录因子在大豆各组织中的表达具有时空特异性,并且可能介导大豆干旱胁迫应答的调控。

To explore the regulatory genes involved in soybean stress response, study focuses on identification and functional analysis of members of the WAX INDUCER1(WIN1/SHN1)family of soybean transcription factors. In this study,using the Arabidopsis AtWIN1 coding sequence as an index and conducting blast comparison in the soybean transcriptome database. A cDNA sequence encoding soybean GmWIN1-1 was obtained. The sequence characteristics of GmWIN1-1 transcription factor and its expression level in various soybean tissues and under drought stress were analyzed by bioinformatics methods and qRT-PCR technology. The results showed that the full length of the sequence was 570 bp encoding 189 amino acids residues. The protein molecular weight and theoretical isoelectric point were 21.2 ku, and 8.45, respectively, and the protein was hydrophilic. The main components of the secondary structure of GmWIN1-1 protein were random coils.The tertiary structure modeling was 45% similar to the amino acid sequence of the template 5wx9.1 protein, which indicating GmWIN1-1 protein was a monomeric protein. Multiple sequence alignment showed that GmWIN1-1 had the highest similarity with AtWIN1. The motif analysis showed that WIN1 protein had 3 motifs, namely: AP2 conservative motif, middle motif, and Cterminal motif. Phylogenetic analysis indicated that GmWIN1-1 was closely related to MtWIN1. The GmWIN1-1 gene promoter contained multiple core cis-elements that respond to stress by PlantCARE software prediction. Real-time fluorescence quantitative PCR analysis showed that the expression of GmWIN1-1 in soybean tissues was significantly different, among which the expression level in flowers was the highest. Under drought stress, the expression level of GmWIN1-1 showed a trend of firstly decreasing slightly, then rapidly increasing and then decreasing. These data suggested that the expression of soybean GmWIN1-1 transcription factor in various soybean tissues was spatiotemporally specific and probably mediated the regulation of response of soybean to drought stress.