作　　者： ; ; ; ;

机构地区： 广东海洋大学水产学院

出　　处： 《中山大学学报（自然科学版）》 2011年第1期110-114,共5页

摘　　要： 将含有新加坡石斑鱼虹彩病毒（Singapore grouper iridovirus,SGIV）ICP18基因的真核表达载体pEGFP-ICP18转染到胖头鲤细胞（Fathead minnow cells,FHM）中进行融合表达,用荧光显微镜观察到ICP18-GFP融合蛋白呈点状分布于FHM细胞的细胞质中。根据SGIV ICP18的序列,设计并体外化学合成了特异性干扰SGIVICP18的siRNA（siRNA-ICP18）,与pEGFP-ICP18共转染到FHM细胞中,通过荧光显微镜观察不同时间点的荧光强度变化。与序列非特异性siRNA（siRNA-negative）阴性对照相比,转染后24~48 h,共转染siRNA-ICP18和pEGFP-ICP18的实验细胞中发荧光的细胞数量较阴性对照少60%~80%左右,说明体外化学合成的siRNA-ICP18可有效抑制FHM细胞中外源导入SGIV ICP18基因的表达。 Singapore grouper iridovirus（SGIV） is a major pathogen resulting in heavy economic losses to grouper aquaculture.In this study,recombant eukaryotic vector pEGFP-ICP18 which inserted with SGIV ICP18 gene was transfected into Fathead minnow（FHM） cells,and ICP18-GFP fusion protein was successfully expressed in FHM cells with a finely punctate cytoplasmic pattern.Candidate siRNA targeting SGIV ICP18 gene（siRNA-ICP18） was designed and chemically synthesized.To investigate the inhibition effect of siRNA-ICP18,pEGFP-ICP18 and siRNA were co-transfected into FHM cells,and the green fluorescence was observed by fluorescence microscope after transfection.The green fluorescence in FHM cells co-transfected with pEGFP-ICP18 and siRNA-ICP18 were 60%~80% fewer than that of negative control,which show the siRNA-ICP18 can effectively silence the extrinsic SGIV ICP18 gene in FHM cells during 24~48 h after transfection.

关 键 词： 干扰 绿色荧光蛋白 石斑鱼虹彩病毒

领　　域： [生物学]