一个烟草葡糖基转移酶基因启动子的克隆与诱导表达

Cloning and Induced Expressing of the Promoter of a Glucosyltransferase-like Gene in Tobacco

利用笔者所在的实验室从烟草中克隆的一个葡糖基转移酶基因（GT-like）,通过PCR扩增得到该基因开放阅读框5′端-1150～0上游调控区。序列分析表明该启动子序列含有多个基因表达调控元件。将GT-like基因启动子与gus报告基因连接,构建植物表达载体pGT-gus,经根癌农杆菌介导法转化W38型烟草。转基因烟草植株GUS组织化学染色结果表明：该基因上游-1150～0序列具有启动子活性,能启动gus基因在烟草叶片中表达,而在根中的表达受时间和环境影响。GUS诱导活性的定量分析结果表明,该启动子的表达不但受甲基茉莉酸的强烈诱导,而且也受水杨酸的强烈诱导。对这一启动子的诱导表达机理的分析将有助于进一步研究甲基茉莉酸和水杨酸之间拮抗作用的相互关系。

The -1 150～0 bp of 5′-upstream regulation region （just upstream ORF） has been amplifled through special PCR from a glucosyitransferase-like（GT-like） gene in tobacco. The inspection of the promoter sequence revealed several boxes important for the regulation of gene expression. The GT-like gene promoter was fused to the 5′-upstream of GUS （β-glucuronidase） coding region in binary vector,designated pGT-gus. The pGT-gus was introduced into W38 tobacco plant by Agrobacterium tumefaciens. The integration of the pGT-gus gene in transgenic tobacco was confirmed by PCR analysis. The resuit of histochemical staining of GUS activity showed that the -1 150～0 bp confer full promoter activity and start up gus gene expression,which was significantly strong in leaf blade and sheath, but the pGT-gus gene expression in root was environmental and temporal differences. Quantification analysis of GUS activity induced showed that the expression of the GT-like gene promoter was induced strongly not only by methyl jasmonate, but also by salicylic acid. The analysis for the expression mechanism of this promoter will redound to the studying on the relationship between the antagonizing salicylic acid and methyl Jasmonate.