摘要
目的:探讨ZEB1基因与肝癌的关系,进一步揭示ZEB1基因在肝癌发生发展过程中的作用。方法:利用基因重组技术构建pCI-neo-ZEB1真核表达载体;脂质体介导转染技术转染肝癌细胞系HepG2。采用Western Blot技术检测重组质粒的转染效率,CCK8比色法测定重组质粒转染对HepG2细胞体外增殖能力的影响,流式细胞术检测重组质粒转染后HepG2细胞凋亡率。结果:重组质粒经Nhel和Xbal双酶切、测序与ZEB1基因序列一致;利用脂质体介导将pCI-neo-ZEB1重组质粒转染HepG2细胞株,经Western Blot技术检测ZEB1基因在蛋白水平稳定高表达;与对照组比较,人HepG2细胞株转染pCI-neo-ZEB1真核表达载体后细胞增殖能力增强,细胞凋亡率明显降低。结论:成功构建重组质粒Pci-neo-ZEB1并转入肝癌细胞株HepG2后,细胞增殖能力增强,凋亡减少。
Objective： To explore the relationship between ZEB1 and hepatocellular carcinoma, and to further reveal the role of ZEB1 in the development of hepatocellular carcinoma. Methods： pCI-neo-ZEB1 eukaryotie expression vector was built by using gene recombination technology, then it was transfected into hepatoma cell line HepG2 cells by liposome. The transfection efficiency was detected by Western Blot. The effects on HepG2 cell proliferation of recombinant plasmids were detected by CCK8 colorimetrie determination. The apoptosis rate of HepG2 cells after plasmid transfection was detected by flow cytometry. Results： The recombinant plasmid was digested by restriction enzymes Nhel and Xbal, and sequenced. The sequencing result was consist- ent with ZEB1 gene sequence in genebank. HepG2 cells transfected with pCI-neo-ZEB1 recombinant plasmids showed a higher expression level of ZEB1 detected by Western Blot. Compared with control group, the proliferation ability of HepG2 cells after transfected with pCI - neo - ZEB1 eukaryotic expression vector was enhanced, and the apoptosis rate was significantly reduced. Conclusion： Recombinant plasmid pCI-neo-ZEBl vector has been successfully constructed and tranfected into HepG2 cells. It provides new insights in understanding the role of ZEB1 gene in the development of hepatocellular carcinoma.
出处
《中西医结合肝病杂志》
CAS
2014年第1期33-36,共4页
Chinese Journal of Integrated Traditional and Western Medicine on Liver Diseases
基金
国家自然科学基金(No.81271808)
关键词
转录因子
ZEB1
真核表达载体
肝细胞癌
transcription factor
ZEB1
eukaryotic expression vector
hepatocellular carcinoma
