CTGF shRNA表达质粒对翼状胬肉成纤维细胞的转染条件优化

目的 用CTGF shRNA表达质粒转染翼状胬肉成纤维细胞,筛选质粒与脂质体的最佳比例,确定最佳转染方案.方法 组织块法培养人眼翼状胬肉成纤维细胞,经细胞免疫组化法(SP法)鉴定,取2~3代细胞,将CTGF shRNA表达质粒与脂质体转染试剂Lipofectamin 2000以不同的比例进行转染,24 h后应用流式细胞仪检测转染效率.测定各组的转染效率,筛选出质粒与脂质体的最适比例.结果 质粒与脂质体的转染比例为1∶2.5时,转染效率最高,达(56.7±1.5)%.结论 本实验为有效地进行体外胬肉成纤维细胞转染及进一步研究重组质粒CTGF shRNA对翼状胬肉干扰奠定了基础.

Optimization of transfection conditions of CTGF shRNA plasmids into pterygium fibroblasts

Objective To investigate the optimal ratio of plasmids to Lipofectamin 2000 after CTGF shRNA plasmids were transfected to pterygium fibroblasts.Methods The primary pterygium fibroblasts were cultured using the method of attachment tissues,and identified with immunohistochemical SP method.Cells of the second or third generation were transfected by using different ratios of CTGF shRNA plasmids to Lipofectamin 2000.After 24 h,the cell transfection efficiency was analyzed by flow cytometry.Then the optimal ratio of plasmids to Lipofectamin 2000 was determined according to the highest transfection efficiency.Results The optimal ratio of plasmids to Lipofectamin 2000 was 1∶2.5,and the transfection efficiency was (56.7±1.5)%.Conclusion This experiment might lay the foundation for performing efficient in vitro transfection into cultured cells and further research on RNA interfere of CTGF gene.