pCDNA3.1-HEGF真核表达质粒转染牙髓干细胞的研究

Study on dental pulp stem cells in transfected with human epidermal growth factor gene in vitro

目的将人表皮生长因子基因转染入牙髓干细胞,并检测其蛋白的表达,为下一步的牙髓干细胞的增殖和分化取得前期的实验基础。方法利用前期实验构建的pcDNA3.1-HEGF,经脂质体转染质粒pcDNA3.1-HEGF入牙髓干细胞,转基因细胞培养48h后作Q-PCR和Western-blot分析。结果成功把pCDNA3.1-HEGF真核表达质粒转染到牙髓干细胞,而且通过检测其表达增强,转hEGF基因细胞经Q-PCR检测,与对照组相比,该基因上调表达了6-21倍,扩增反应产物溶解温度较均一,目的基因具有很好的特异性,反应体系良好,Western-blot检测到HEGF表达明显升高。结论质粒pcDNA3.1-HEGF在脂质体介导下成功转染牙髓干细胞,而且表达增强。

Objective To transfect the human epidermal growth factor gene into the dental pulp stem cells,and detect the expression of the protein,so as to achieve early experimental basis for the proliferation and differentiation of the dental pulp stem cells.Methods Using PcDNA3.1-HEGF built in a previous experiment,the liposometransfection plasmid pcDNA3.1-HEGF was transfected into dental pulp stem cells,mediated by Lipefectin2000.After being cultured for 48 hours,the transcription and expression of the HEGF gene in transfected keratinocytes were investigated by Q-PCR and Western-blot.Results The pCDNA3.1-HEGF eukaryotic expression plasmid was successfully transfected into dental pulp stem cells,and enhanced the gene expression 6-12 times compared with that of the control group.The solution temperature of amplificate reaction product was uniform,the purpose gene showed good specificity,and the reaction system was good.Conclusion The plasmid pcDNA3.1-HEGF mediated by the liposome was successfully transfected into dental pulp stem cells and enhanced the gene expression.