快速检测干旱和脱水可诱导植物启动子瞬间表达特性的方法

Rapid Determination of Transient Expression Characteristics of Drought-and Dehydration-Inducible Promoters from Plants

选择合适的诱导表达启动子是开展植物耐干旱和脱水等非生物逆境转基因研究的重要环节。我们通过几年的研究,已建立了一套以大麦幼苗完整活体和植物离体叶片为主要材料通过瞬间表达鉴定来快速检测干旱和脱水可诱导基因启动子表达特性的方法。来自大麦和水稻的启动子Dhn4 s、Dhn8 s、HVA1 s、Rab16B j、wsi18 j在大麦、小麦、水稻、高粱和蕨类植物的离体叶片中经干燥诱导可以瞬间表达GFP,在绿豆、番茄叶片中不表达。鉴定了HVA1 s和wsi18 j在大麦不同器官或组织中启动子的定性表达情况。进一步建立了GFP荧光点/GUS染色点计数分析和GUS活性/XYN活性测定分析的启动子表达的定量分析方法,并讨论该方法在环境可诱导植物启动子功能分析中的应用价值和前景。

The selection of suitable inducible-promoters is one of the most important chains in transgenic researches for plant tolerances to drought, dehydration and other abiotic stresses. Based on laboratory researches in the past years, it was set up a set of methods that could be used to rapidly determine the transient expression characteristics of drought-and dehydration-inducible promoters by using whole barley seedling and plant leaves in vitro as bombardment objects. Promoters Dhn4s, Dhn8s, HVAls, Rab16Bj and wsi18j isolated from barley and dce were able to express GFP in the leaves of barley, wheat, rice, sorghum and fern after drying treatment, but not in laves of mungbean and tomato. The qualitative expression of HVA1s and wsi18j in different organs and tissues of barley was identified. A method for quantitative analysis of promoter transient expression was established by means of GFP foci/GUS foci counting or GUS activity/XYN activity test. Finally, the value and prospect of the methodology were discussed in terms of its application to the analysis of plant promoters inducible by environmental factors.