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目的 研究p16基因对肿瘤细胞生长抑制及细胞周期阻滞作用。方法 将p16cDNA插入逆转录病毒载体pLXSN构建成p16基因逆转录病毒重组体pLp16SN。利用基因转染方法 ,将pLp16SN及pLXSN导入逆转录病毒包装细胞系PA317细胞 ,获得逆转录病毒。用逆转录病毒感染Bcap 37乳腺癌细胞 ,经G4 18筛选获阳性克隆。利用Northern和Westernblotting方法检测p16基因的表达。检测转基因细胞的生长速度 ,细胞周期及裸鼠成瘤等细胞生物学行为的改变。结果 Northern及Westernblotting显示转染p16基因的Bcap 37细胞p16基因mRNA及蛋白质表达明显增高。此细胞较未转染基因的Bcap 37细胞和转染空载体的Bcap 37细胞生长速度慢 ,G1期细胞比率增高 ,裸鼠接种成瘤体积小。结论 p16基因高表达能够抑制乳腺癌细胞Bcap 37的生长 ,并阻滞细胞从G1期进入S期
Objective To study the effect of p16 gene on tumor cell growth inhibition and cell cycle arrest. Methods p16 cDNA was inserted into retroviral vector pLXSN to construct p16 gene retrovirus recombinant pLp16SN. Using gene transfection method, pLp16SN and pLXSN were introduced into retrovirus packaging cell line PA317 to obtain retrovirus. Bcap 37 breast cancer cells were infected with retrovirus and positive clones were screened by G418. The expression of p16 gene was detected by Northern and Western blotting. Detection of the growth of transgenic cells, cell cycle and tumorigenesis of nude mice and other biological behavior changes. Results Northern and Western blotting showed that the expression of p16 mRNA and protein in Bcap 37 cells transfected with p16 gene was significantly increased. Compared with Bcap 37 cells transfected with empty vector and Bcap 37 cells transfected with empty vector, the cell growth rate was slower than that of untransfected cells, the ratio of cells in G1 phase increased, and the inoculation volume of nude mice was small. Conclusion The overexpression of p16 gene can inhibit the growth of Bcap 37 breast cancer cells and arrest the cells from G1 phase to S phase