摘要
目的:制备在人大肠癌细胞系LoVo中稳定抑制PLCγ1(phospholiase C-gamma 1)的重组病毒,建立PLCγ1表达降低的细胞系,以便客观研究该基因的作用。方法:制备产生PLγ1 siRNA分子的重组慢病毒,在慢病毒感染LoVo细胞后以杀稻瘟菌素筛选稳定表达细胞系。应用Western-blot和RT-PCR对细胞中PLCγ1的蛋白和mRNA表达水平进行分析,XTT法和黏附能力测定分别检测其对细胞增殖和黏附的影响。结果:PLCγ1 siRNA显著下调了LoVo细胞中PLCγ1的表达,所构建的重组慢病毒导入的siRNA有较高的沉默效率,LoVo细胞的黏附能力显著降低,细胞增殖无明显改变。结论:研究证实了 PLCγ1基因可显著地影响大肠癌LoVo细胞的黏附能力,为利用PLCγ1介导的信号通路进行肿瘤基因治疗提供了实验依据。
Objective： To preare recombinant lentivirus stably suppressing PLCγ1 in human colorectal carcinomas LoVo celis, so as to establish LoVo cell lines deficient in PLCγ1 and to investigate the role of this gene. Methods：Reeombinant lentivirus produeeing PLCγ1 siRNA were prepared After LoVo cells were transduced with lentivirust, stably transduced cells were selected by Blasticidin. The protein and mRNA expression of PLCγ1 was examined by Western-blot and RT-PCR analysis. The effect of the lentivirus on the cell proliferation and cell adhesion was analyzed by XTT method and cell adhesion assay, respectively. Results： PLCγ1 siRNA knocked down PLCγ1 expression in LoVo cells obviously. The silenced efficiency of siRNA transducted by recombinant lentivirus was very high. Adhesion of human colorectal carcinomas LoVo cell Lines was significantly decreased, while proliferation was not affected. Conclusion： Our research confirm that PLCγ1 plays an important role in cell adhesion of colorectal carcinomas, and provides experimental evidences for targeting PLCγ1 in gene therapy against cancer.
出处
《中国肿瘤生物治疗杂志》
CAS
CSCD
2006年第1期18-21,共4页
Chinese Journal of Cancer Biotherapy
基金
国家自然科学基金(39870381)广东省自然科学基金(020015)广东省自然科学重点科学基金(05102580)
