核盘菌一分支酸变位酶同源效应蛋白的亚细胞定位

前期研究表明核盘菌Sclerotinia sclerotiorum产生的一种分支酸变位酶同源效应蛋白能够提高其致病能力。为了进一步研究该效应蛋白在核盘菌致病过程中的作用机制,我们通过构建GFP融合蛋白对其分泌行为和亚细胞定位开展了研究。首先通过PCR技术扩增了该基因的启动区+信号肽(SP)+叶绿体定位肽(CTP)的DNA序列,构建GFP融合载体,然后借助REMI技术转化核盘菌原生质体,筛选GFP能够表达的转化子,最后接种转化子菌丝于烟草叶片,激光共聚焦检测GFP荧光信号,发现GFP荧光与叶绿体自体荧光共定位,表明该效应蛋白可分泌进入寄主叶绿体内发挥作用。

Subcellular localization of an effector homologous with chorismatemutase in Sclerotinia sclerotiorum

It has been proved that the effector homologous with chorismate mutase was related with pathogenicity of Sclerotinia sclerotiorum. In order to explore the pathogenicity mechanism of the effector, we analyzed the secretory and subcellular localization through constructing the GFP fusion protein. The DNA sequence of promoter +SP+CTP was cloned by PCR. Then the vector of GFP fusion promoter +SP+CTP was constructed, transformed in the protoplast of S.sclerotiorum using the REMI. The transformants in which GFP could express normally were obtained and the transformants mycelia were inoculated on the leaves of tobacco and the fluorescence signals were detected through confocal microscope. As a result, it was found that the GFP fusion protein could be secreted into host cell and colocalized with the chloroplast. The result indicated that the effector may play its role in the host chloroplasts.