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目的:通过稳定、高效、低毒的基因转染方法将nm23-H1基因导入该基因缺失的人肺大细胞癌株L9981,从基因及蛋白水平鉴定nm23-H1基因转染肺癌细胞的建立。方法:以JM109为宿主菌,转化质粒载体PCMV-neo-Bam-nm23-H1,制备nm23-H1基因的真核表达载体;利用阳离子脂质体Lipofectamine介导,将nm23-H1基因导入L9981细胞株。采用RT-PCR技术检测转染后nm23-H1基因的表达,并用Western blot技术检测转染后nm23-H1蛋白的表达。结果:转染后的L9981细胞株中可检测到nm23-H1 mRNA及nm23-H1蛋白的阳性表达。结论:建立表达nm23-H1基因的肺癌细胞株L9981,为研究nm23-H1对肺癌恶性转移表型的逆转作用作基础准备。
OBJECTIVE: To construct nm23-H1 gene transfected lung cancer cell line nm23-H1 by genetically and stably, efficiently and with low toxicity gene transfer into human lung cancer cell line L9981. Methods: The eukaryotic expression vector of nm23-H1 gene was constructed by transforming plasmid vector PCMV-neo-Bam-nm23-H1 with JM109 as the host strain. Lipofectamine was used to transduce the nm23-H1 gene into L9981 cell line . The expression of nm23-H1 gene was detected by RT-PCR and the expression of nm23-H1 protein was detected by Western blot. Results: The positive expression of nm23-H1 mRNA and nm23-H1 protein in transfected L9981 cells was detected. CONCLUSION: The lung cancer cell line L9981 expressing nm23-H1 gene was established to study the reverse effect of nm23-H1 on the malignant metastasis of lung cancer.