鸭梨PPO与GFP融合基因的烟草转化及亚细胞定位观察

将鸭梨PPO基因与绿色萤光蛋白GFP基因相融合共同进行遗传转化的方式,对鸭梨多酚氧化酶开展细胞定位研究。通过克隆该酶基因除终止密码子TAA外长度为1 779bp的CDS序列,与绿色荧光蛋白基因重组构建了荧光表达载体pBI121-PPO-GFP,借助农杆菌转化烟草,转基因烟草叶片细胞经激光扫描共聚焦显微镜观察,绿色荧光蛋白荧光与叶绿体自发荧光相重合。结果表明鸭梨多酚氧化酶为叶绿体蛋白质。

Transformation of tobacco plants by Yali PPO-GFP fusion gene and observation of subcellular localization

To explore the subcellular localization of Polyphenol oxidase(PPO)from Pyrus bretschneideri,the 1 779 bp cDNA of PPO gene was cloned and fused with GFP in frame to construct a binary vector pBI121-PPO-GFP. Then,the binary vector was transformed into Nicotiana tabacum by the tumefanciens-mediated method. Using confocal laser scanning microscopy,green fluorescent signals were localized in chloroplasts of the transformed Nicotiana tabacum cell. This results suggested that the Polyphenol oxidase from Pyrus bretschneideri was a chloroplast protein.