杂交构树UDP-葡萄糖脱氢酶基因编码蛋白的亚细胞定位及其启动子5'端缺失片段的功能分析

Subcellular Localization of the Protein Coded by the UDP-Glucose Dehydrogenase Gene from Paper Mulberry and Functional of Its Promoter 5′-end Deletion Fragment

为了研究杂交构树UDP-葡萄糖脱氢酶基因(DDBJ,BpUGDH基因登录号为LC457701)启动子不同区域的表达活性,利用5′端缺失及同源重组实验技术,将5个不同长度的BpUGDH启动子5′端缺失片段与GUS基因连接,并通过农杆菌介导法瞬时转化烟草;同时,为了定位BpUGDH基因编码的蛋白在细胞中表达的具体位置,利用GFP报告基因融合目的基因进行蛋白质的亚细胞定位。结果显示:BpUGDH基因启动子-244 bp以内的序列均能介导GUS基因的诱导表达,并且-973、-465、-355、-281和-244 bp之间的区域可能对BpUGDH基因启动子的活性发挥着至关重要的作用。另外,BpUGDH基因编码蛋白的亚细胞定位结果显示:BpUGDH位于叶绿体中。

We analyzed the function of cis-acting elements of UDP-Glucose Dehydrogenase Gene from Paper Mulberry(DDBJ,BpUGDH accession No.LC457701)promoter,the five different lengths of BpUGDH promoter 5′-terminal deletion fragment were ligated with the GUS gene by using 5′terminal deletion and homologous recombination techniques,and conducted through heterologous expression in Nicotiana benthamiana by agroinfiltration method.In order to locate the specific location of the protein encoded by BpUGDH gene in cells,subcellular localization of BpUGDH protein was carried out by using the fusion target gene of GFP reporter gene.The results show that the sequences within-244 bp can mediate the induction of GUS gene expression,and the region between-973,-465,-355,-281,-244 bp of the BpUGDH promoter were crucial for regulating the promoter activity.Furthermore,BpUGDH was located in chloroplast.