烟草DREB转录因子新基因的克隆与功能分析

Cloning and functional analysis of a novel DREB transcription factor from Nicotiana benthamiana

采用同源克隆及cDNA末端快速扩增法(rapid amplification of cDNA ends,RACE),从烟草中克隆到1个新的烟草DREB基因,命名为NbDREB2 a.序列分析表明该基因全长1191 bp,编码330个氨基酸,具有典型的DREB转录因子保守的AP2结构域.实时RT-PCR分析表明,该基因能在烟草的根、茎、叶中表达,受低温、干旱和高盐诱导.酵母转录激活实验证明,该基因可以特异性结合DRE顺式作用元件,并能激活报告基因的表达.为了研究该基因在植物抗逆反应中的功能,利用农杆菌介导法,将该基因转化到模式生物拟南芥中,筛选到了单拷贝插入的、纯合的拟南芥株系;对获得的转基因拟南芥进行干旱及高盐处理,结果表明,与对照相比,超量表达该基因能够明显地提高拟南芥对干旱及高盐的抗性.

A novel dehydration responsive element binding protein（DREB） transcription factor gene named NbDREB2a was isolated from Nicotiana benthamiana using homology-based and RACE technology.The full-length of the NbDREB2a cDNA was 1 191 bp and was predicted to encode a protein of 330 amino acids with a conserved AP2 domain.Quantitative real-time RT-PCR analysis revealed that NbDREB2a was expressed ubiquitously in tobacco including leaf,root and stem and could be induced by drought,low temperature and high salt stresses.NbDREB2a protein could bind to the DRE element and activate the expression of downstream reporter genes in yeast cells.By using Agrobacterium-mediated transformation,the NbDREB2a gene was successfully introduced into Arabidopsis.Transgenic Arabidopsis with single copy of NbDREB2a was obtained for further research.The results showed that overexpression of NbDREB2a enhanced tolerance to drought and high salt stresses in transgenic Arabidopsis.