【摘要】 目的:构建带Flag标签的人转录因子激活蛋白2γ（TFAP₂γ）基因真核表达载体,获得Flag-TFAP₂γ蛋白,检测其对乳腺癌细胞生长的影响。方法:采用PCR技术从乳腺文库中扩增出人TFAP₂γ基因,并将其正确插入Flag载体;将重组质粒与空载体转染人乳腺癌细胞系ZR75-1和MCF-7细胞后,Western印迹检测表达情况,并进行生长曲线实验。结果:双酶切和测序鉴定表明,Flag-TFAP₂γ真核表达质粒构建成功,转染ZR75-1和MCF-7细胞后获得表达;生长曲线实验结果表明,TFAP₂γ可促进乳腺癌细胞的生长。结论:构建了带Flag标签的人TFAP₂γ基因真核表达载体,并能在乳腺癌细胞ZR75-1和MCF-7中表达,且能促进细胞的生长。本实验为进一步研究TFAP₂γ在乳腺癌中的功能奠定了基础。更多还原

【Abstract】 Objective:To construct the eukaryotic expression vector of human transcription factor activator protein 2 gamma（TFAP₂γ） gene labelled with FLAG tag and detect its activity.Methods:Human TFAP₂γ coding region was amplified from human mammary cDNA library by PCR and cloned into FLAG vector.Human ZR75-1cells and MCF-7 cells were transfected with the recombinant plasmid FLAG-TFAP₂γ and the expression was detected by Western blotting.The growth curves of the breast cancer cells transfected with FLAG-TFAP₂γ and empty vector were performed to analyze the effect of TFAP₂γ on breast cancer cell proliferation.Results:TFAP₂γ eukaryotic expression vector labeled with FLAG was verified by double digestion and DNA sequencing.Human TFAP₂γ protein was expressed in human ZR75-1 cells and MCF-7 cells as was shown by Western blotting.ZR75-1 cells and MCF-7 cells transfected with FLAG-TFAP₂γ grew faster than those transfected with empty vector.Conclusion:FLAG-TFAP₂γ promotes proliferation of ZR75-1 cells and MCF-7 cells.The whole work laid the foundation for the further study on TFAP₂γ function in breast cancer.更多还原