纤毛鹅观草DREB类基因RcDREB1的克隆及其蛋白结合干旱应答元件DRE的功能验证

Cloning of a DREB-like Gene RcDREB1 in Roegneria ciliar and Functional Identification of its Protein Binding Specificity to the Dehydration-responsive Element DRE

以纤毛鹅观草为材料,通过RT-PCR和RACE技术,首次克隆了一个纤毛鹅观草DREB类基因的全长cDNA序列,命名为RcDREB1。该基因编码蛋白含有一个保守的AP2结构域,表明该基因是AP2大家族的一个新成员。种系发生树分析表明,RcDREB1与小麦AP2家族DREB类基因高度同源。酵母单杂交试验表明,RcDREB1表达的蛋白具有特异结合干旱应答DRE元件的功能;通过实时定量PCR分析发现,高盐、干旱、重金属镉和低温胁迫均可诱导RcDREB1表达,但其对高盐反应较为显著。本研究表明,RcDREB1是DREB类转录因子基因,在介导植物响应逆境信号方面可能发挥着重要生物学功能,为进一步分析小麦族的进化和转基因应用奠定了基础。

In this paper, a full-length cDNA of a DREB-like gene was cloned firstly from Roegneria ciliaris through RT-PCR and RACE technologies,and named as RcDREB1. RcDREB1 contained a conserved AP2 domain, suggesting that this gene was a new transcription factor member in the AP2 superfamily. Phylogenetic tree analysis showed that RcDREB1 shared a high similarity with the DREB-like genes in Triticum aestivum. Through yeast one- hybrid experiment,RcDREB1 was verified to possess an ability to specifically bind the drought-responsive DRE ele- ment. Using real-time quantitative PCR, we found that all of the stresses of high salt, drought, heavy metal （CdC12 ）, and low temperature could up-regulate RcDREB1 expression, but only high salt showed more strong induction of RcDREB1. These results suggested that RcDREB1 was a novel DREB type transcription factor gene and might play important roles in mediating the transduction of distinct signaling cues initiated by those of adverse environmental factors. This DREB-like gene could be useful in elucidating the evolution pathway of Triticeae and in the application of transgenic wheat breeding.