新城疫病毒感染细胞的双向凝胶电泳方法建立及其初步分析

Establishment of Two-dimensional Gel Electrophoresis for Newcastle Disease Virus Infected-cells and Its Preliminary Application

为了研究新城疫病毒(NDV)和宿主细胞之间的相互作用关系,本研究以NDV感染鸡胚成纤维细胞(CEF)为材料,对细胞培养方式、样品制备方法、上样量、等电聚焦以及凝胶染色等步骤进行一系列优化,建立了NDV感染CEF的双向凝胶电泳技术。运用PDQuest8.0.1软件对电泳图谱分析后显示:17cmIPG胶条(pH4～7)对应的凝胶上可检测到800个左右蛋白点、蛋白点匹配率达90%,表明NDV感染CEF蛋白质组双向凝胶电泳模型稳定、分辨率高、重复性好。电泳图谱上共发现36个差异表达明显的蛋白点,其中上调表达蛋白点有16个,下调表达蛋白点15个,新出现蛋白点3个,消失蛋白点为2个。NDV感染CEF双向凝胶电泳技术的建立为NDV的致病机理研究提供了有力的工具。

To establish a stable technique of two-dimensional gel electrophoresis （2-DE）for chicken embryo fibroblasts （CEF） infected with Newcastle disease virus （NDV）,eett culture,sample preparation,loading capacity,isoelectric focusing, staining and other steps were optimized using NDV-infected CEF as model system. About 800 protein spots on 17 cm IPG strip （pH 4 to 7） stained with blue sliver were analyzed by the software PDQuest8.0.1 ,and the match rate of protein spots was over 90%. The analysis of multiple 2-DE gets revealed that a total of 36 protein spots differentially expressed during NDV infection,including 16 up-regulated protein spots,15 down-regulated protein spots,3 new emerging protein spots and 2 disappear protein spots. Those differentially expressed protein spots were useful to further investigation of the interaction between NDV and its hosts.