中华实验外科杂志
中華實驗外科雜誌
중화실험외과잡지
CHINESE JOURNAL OF EXPERIMENTAL SURGERY
2014年
1期
108-110
,共3页
刘岳%陈谦学%田道锋%刘宝辉%吴立权
劉嶽%陳謙學%田道鋒%劉寶輝%吳立權
류악%진겸학%전도봉%류보휘%오립권
B细胞淋巴瘤/白血病-2%胶质瘤%质粒
B細胞淋巴瘤/白血病-2%膠質瘤%質粒
B세포림파류/백혈병-2%효질류%질립
B lymphocytes/leukemia-2%Glioma%Plasmid
目的 构建人B细胞淋巴瘤/白血病-2(bcl-2)基因质粒体并检测其在胶质瘤细胞株U251内的表达.方法 构建PEGFP-bcl-2质粒体,应用FUGENE HD转染质粒体进入U251,G418筛选,挑出单克隆扩增,建立稳定细胞株N1-U251和bcl-2-U251.将实验分为:空白组(U251细胞)、N1组(N1-U251)、bcl-2组(bcl-2-U251),检测bcl-2基因和蛋白表达；使用噻唑蓝(MTT)法检测N1组(N1-U251),bcl-2组细胞活性.结果 PEGFP-bcl-2质粒体构建成功,N1-U251、bcl-2-U251稳定细胞株构建成功,在空白组、N1组、bcl-2组bcl-2蛋白表达量分别为:0.012±0.003、0.011 ±0.003、0.115±0.025.细胞活性分别为:1.015±0.005、1.120±0.004、3.312±0.004,bcl-2组与对照组比较差异有统计学意义(P＜0.05).结论 成功构建人bcl-2质粒体并建立高表达bcl-2的细胞株,高表达bcl-2可以提高U251细胞活性.
目的 構建人B細胞淋巴瘤/白血病-2(bcl-2)基因質粒體併檢測其在膠質瘤細胞株U251內的錶達.方法 構建PEGFP-bcl-2質粒體,應用FUGENE HD轉染質粒體進入U251,G418篩選,挑齣單剋隆擴增,建立穩定細胞株N1-U251和bcl-2-U251.將實驗分為:空白組(U251細胞)、N1組(N1-U251)、bcl-2組(bcl-2-U251),檢測bcl-2基因和蛋白錶達；使用噻唑藍(MTT)法檢測N1組(N1-U251),bcl-2組細胞活性.結果 PEGFP-bcl-2質粒體構建成功,N1-U251、bcl-2-U251穩定細胞株構建成功,在空白組、N1組、bcl-2組bcl-2蛋白錶達量分彆為:0.012±0.003、0.011 ±0.003、0.115±0.025.細胞活性分彆為:1.015±0.005、1.120±0.004、3.312±0.004,bcl-2組與對照組比較差異有統計學意義(P＜0.05).結論 成功構建人bcl-2質粒體併建立高錶達bcl-2的細胞株,高錶達bcl-2可以提高U251細胞活性.
목적 구건인B세포림파류/백혈병-2(bcl-2)기인질립체병검측기재효질류세포주U251내적표체.방법 구건PEGFP-bcl-2질립체,응용FUGENE HD전염질립체진입U251,G418사선,도출단극륭확증,건립은정세포주N1-U251화bcl-2-U251.장실험분위:공백조(U251세포)、N1조(N1-U251)、bcl-2조(bcl-2-U251),검측bcl-2기인화단백표체；사용새서람(MTT)법검측N1조(N1-U251),bcl-2조세포활성.결과 PEGFP-bcl-2질립체구건성공,N1-U251、bcl-2-U251은정세포주구건성공,재공백조、N1조、bcl-2조bcl-2단백표체량분별위:0.012±0.003、0.011 ±0.003、0.115±0.025.세포활성분별위:1.015±0.005、1.120±0.004、3.312±0.004,bcl-2조여대조조비교차이유통계학의의(P＜0.05).결론 성공구건인bcl-2질립체병건립고표체bcl-2적세포주,고표체bcl-2가이제고U251세포활성.
Objective To construct the plasmid of B lymphocytes/leukemia-2 (bcl-2) and observe its expression in U251 cells.Methods Plasmid of bcl-2 was constructed and transfected into U251 cells by FUGENE HD.G418 was used to kill the cells that were transfected unsucessfully.Western blotting and reverse transcriptase polymerase chain reaction (RT-PCR) were used to observe the expression of bcl-2.Three groups were set up:control group,N1 group and bcl-2 group.Methyl thiazol tetrazolium (MTT) assay was used to measure the viability of U251 cells in N1 group and bcl-2 group.Results The plasmid PEGFPbcl-2 and bcl-2-U251 cells were constructed successfully.The expression of bcl-2 in control group,N1 group and bcl-2 group was 0.012 ±0.003,0.011 ±0.003 and 0.115 ±0.025,and the viability of U251 cells was 1.015 ±0.005,1.120 ± 0.004 and 3.312 ± 0.004,respectively.There was significant difference between bcl-2 group and control group (P ＜ 0.05).Conclusion The plasmid PEGFP-bcl-2 was constructed successfully and the overexpression system is also established,and bcl-2 overexpression can increase the viability of U251 cells.
