摘要
目的构建抑癌基因p27高表达细胞株,研究其对子宫颈癌HeLa细胞增殖的影响。方法运用RT-PCR技术从HeLaRNA中扩增p27 cDNA,连接酶连接至真核表达载体pcDNA3.1(+),所得的重组质粒pcDNA3.1(+)-p27用脂质体法转染子宫颈癌HeLa细胞,使用RT-PCR以及Western blot法筛选p27基因高表达的细胞株。MTT法和流式细胞术检测p27基因高表达对细胞增殖的影响。结果成功获得p27高表达的HeLa细胞株。细胞增殖分析和流式细胞术结果显示,p27基因高表达对细胞增殖有抑制作用,并明显增加G1期的细胞数(由44.4%增加到59%,P<0.05)。结论 p27基因在HeLa细胞中高表达能通过抑制细胞G1/S转换从而抑制细胞增殖,提示p27基因可能成为子宫颈癌基因治疗的靶基因。
Purpose To study the effect of p27 on the growth of HeLa cells by constructing a p27 overexpression cell line. Methods p27 cDNA was amplified by RT-PCR and then cloned into pcDNA3.1 ＋ vector. The resulted plasmid pcDNA3.1 （ ＋ ） -p27 was trans- fected into HeLa cells by lipofectinamin reagent. RT-PCR and Western blot were used to select single clones with p27 oerexpression. Effects of p27 on cell growth and cell cycle were detected by MTT method and flow cytometry assay. Results The HeLa cell lines with p27 overexpression were obtained successfully. Overexpression of p27 inhibited cell growth and caused G1/S arrest with increased G1 population （ from 44. 4% to 59% ）. Conclusion Overexpression of p27 gene can inhibit HeLa cell growth by interfering the cell ey- cles, suggesting that p27 may be a good candidate for gene therapy of carcinoma in cervix.
出处
《临床与实验病理学杂志》
CAS
CSCD
北大核心
2012年第6期631-635,共5页
Chinese Journal of Clinical and Experimental Pathology
基金
国家自然科学基金(30873282)
湖北省卫生厅科研项目(NX2011-9)
