稳定表达siRNA-NUAK1基因95D细胞系的建立

目的建立稳定表达siRNA-NUAK1基因的人肺巨细胞高转移株95D细胞系,为研究NUAK1在肺癌细胞转移侵袭中的作用机制提供基础。方法利用脂质体把pGCsilencerU6/GFP/Neo-RNAi-NUAK1重组质粒转染95D肺癌细胞,G418筛选稳定转染细胞系,从中挑出单克隆,分别通过荧光显微镜和RT-PCR法鉴定单克隆细胞和NUAK1基因的转染、抑制效果。结果从稳定转染细胞中,成功挑出单细胞克隆3株,其中2株细胞的NUAK1基因的表达被显著抑制。结论利用siRNA技术成功建立了稳定表达siRNA-NUAK1基因的95D细胞系,为研究NUAK1因子与肺癌转移侵袭的关系及机制提供了细胞模型。

Establishment of a stable siRNA-NUAK1 95D cell line

Objective To establish 95D cell lines that stably express siRNA-NUAK1 for the study of the relationship between NUAK1 expression and the metastasis invasion of lung tumor cell.Methods The plasmid of pGCsilencerU6/GFP/Neo-RNAi-NUAK1 were used to transfect 95D cell with lipofectamine.Stable transfection clone cells were selected out by G418,the effects of transfection and NUAK1 gene silencing were identified respectively by fluorescence microscopy and RT-PCR.Results Three clones were picked out from stable transfection cells,and two clones NUAK1 gene expression was significantly inhibited.Conclusion We successfully selected out 95D cell line expressed siRNA-NUAK1 gene stably and efficiently in this study.This work has provided a cell model for investigating the relationship between the NUAK1 and the metastasis invasion of lung cancer cell malignancy.