过量表达菊花DmDREBa基因提高转化烟草耐低温能力

Overexpression of DmDREBa Gene Significantly Enhances Low Temperature Tolerance in Transgenic Tobacco

各种环境因素,如干旱、高盐、激素和低/高温等非生物胁迫对植物的生长发育造成很大影响。DREB转录因子在植物抵抗非生物胁迫中起到关键作用。本研究通过根癌农杆菌介导的叶盘转化法将菊花DmD REBa基因导入烟草中并进行了耐低温能力分析。研究利用PCR的方法鉴定出了43株转基因阳性植株。随机选取其中9株转基因植株,有7株在RNA转录水平能够表达。Southern杂交检测表明,DmD REBa基因以1~3个拷贝形式随机插入到烟草基因组中。胁迫处理结果表明,DmD REBa基因明显增强了转基因烟草抵抗低温能力。通过叶片上下表皮气孔密度检测,发现转基因烟草的蒸腾失水量远远低于对照野生型。进一步对低温胁迫下转基因烟草的丙二醛含量进行测定分析,发现转基因烟草丙二醛含量比野生型烟草低22.29%。综上结果表明,DmD REBa基因能够提高转基因烟草对低温的耐受能力,为菊花DREB转录因子的深入研究提供理论依据,并为进一步解析菊花DREB基因功能奠定基础。

Many environmental factors such as drought, salt stress, hormones and low/high temperature influence the growth and development of plants. It has been found that DREB transcription factors play important roles in plants against different abiotic stresses. In this study, DmDREBa （GenBank accession No. EF490996） , a DREB-like gene isolated from chrysanthemum was overexpressed in tobacco plants by mediation of Agrobacterium tumefaciens under the control of the constitutive promoter CaMV 35S, and 43 transgenic tobacco plants were obtained by PCR method. 9 transgenic tobacco plants were randomly selected to perform RT-PCR investigation and 7 plants indicated RNA transcripts accumulation. The southern blot result showed that 1 -3 copies of DmDREBa were randomly inserted. The tolerance of transgenie tobacco plants was increased comparing with the wild type tobacco plants under low temperature condition. The water loss of transgenic tobacco was less than wild type plants after natural transpiration. Moreover, the density of the stomata in the epidermis was observed. The result showed that the numbers of stomata in upper or lower epidermis of transgenic plants were reduced comparing to wild type tobacco. The result of t test demonstrated that the differences b6tween the upper and lower epidermis of transgenic plants and wild type were significant. Furthermore, the con^nt of malondialdehyde （MDA） in transgenic plants was lower than that in wild type tobacco plants at lower temperature condition, which indicated the tolerance ability of transgenic plants was increased. All these results indicated that DmDREBa improved tolerance ability of trangenic tobacco plants to the lower temperature stress. This research will provide a very useful reference for functional analysis of DREB genes in chrysanthemum in the future.