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目的从人肝癌细胞系HepG2中获得PinX1基因,构建真核表达载体,转染Hek293细胞。方法采用RT-PCR技术从HepG2中扩增PinX1,克隆入真核表达载体pcDNA3,重组质粒转染Hek293细胞,免疫组化法检测蛋白的表达。结果RT-PCR方法扩增获得PinX1基因,构建真核表达载体pcDNA3-PinXl-vsv重组质粒转染Hek293细胞,免疫组化检测结果表明在细胞核内有PinX1表达。结论成功获得PinX1基因,构建的重组载体可在Hek293细胞内表达,为探索PinX1在端粒、端粒酶调控中的作用机制奠定基础。
Objective To obtain PinX1 gene from human hepatocellular carcinoma cell line HepG2, construct eukaryotic expression vector and transfect Hek293 cells. Methods PinX1 was amplified from HepG2 by RT-PCR, cloned into eukaryotic expression vector pcDNA3, and transfected into Hek293 cells. The protein expression was detected by immunohistochemistry. Results The PinX1 gene was amplified by RT-PCR. The recombinant plasmid pcDNA3-PinXl-vsv was constructed and transfected into Hek293 cells. Immunohistochemistry results showed that PinX1 was expressed in the nucleus. Conclusion The PinX1 gene was successfully obtained and the recombinant vector was constructed and could be expressed in Hek293 cells. This study may provide a basis for exploring the role of PinX1 in telomere and telomerase regulation.