杂交鹅掌楸LhARF2基因启动子的克隆及瞬时表达分析

Cloning and Transient Expression Analysis of LhARF2 Promoter in Liriodendron Hybrids

生长素响应因子(Auxin response factor,ARF)参与调控植物生长发育的多个途径。本研究利用Tail-PCR技术从杂交鹅掌楸中克隆LhARF2基因5'端上游2637bp序列,即pLhARF2启动子,并利用生物信息学软件PlantCARE分析其包含的调控元件,发现其含有启动子必需元件TATA-box和CAAT-box,以及压力响应、激素响应、光信号转导和代谢循环过程元件。构建pLhARF2-pCAMBIA1300:GUS植物表达载体并瞬时浸染本氏烟草叶片,利用GUS组织化学染色法鉴定烟草叶片中瞬时表达活性,发现在烟草叶片中有蓝色斑块,但颜色较浅。实验结果推测杂交鹅掌楸pLhARF2启动子可以调控GUS基因活性,为下一步研究启动子遗传转化的组织表达活性奠定了基础。

Auxin response factor ( ARF) participates regulating plant growth and development in multiple pathways.This study using the Tail - PCR technology to cloning LhARF2 gene of upstream sequence from the Liriodendron hybrids,the length was 2 637 bp,named pLhARF2 promoter,and using bioinformatics software PlantCARE analysis regulatory elements,containing the promoter required components TATA box and CAAT box,and pressure response,hormone response,light signal transduction and the metabolic cycle components.pLhARF2 - pCAMBIA1300: GUS plant expression vector was constructed and immediately impregnated into tobacco leaves.GUS histochemical stain method was used to identify the transient expression activity in tobacco leaves.It was found that there were blue patches in tobacco leaves,but the color was relatively light.The result speculate that the pLhARF2 promoter had regulatory activity of GUS gene from Liriodendron hybrids, which laid a foundation for the further study of the promoter tissue expression activity in genetic transformation.