摘要
目的构建重组基因表达载体PcDNA-sTRAIL并观察其对C6胶质瘤细胞的抑制作用。方法通过PCR扩增和定向克隆技术构建重组真核表达载体PcDNA-sTRAIL。以阳离子脂质体介导转染C6胶质瘤细胞并用流式细胞仪观测对细胞增殖的影响。结果经过测序鉴定和蛋白表达检测证实,重组真核表达载体PcDNA-sTRAIL构建成功。阳离子脂质体介导质粒转染C6胶质瘤细胞阳性率达50%。脂质体PcDNA-sTRAIL转染C6细胞后,相比未转染、空质粒转染、Lipofectin转染组C6胶质瘤细胞增殖抑制、细胞凋亡明显增加(t分别=9.52、7.20、2.84,P均<0.05)。结论成功构建了sTRAIL基因的真核表达载体,并从体外实验中初步证实了其对C6胶质瘤细胞增殖具有明显的抑制作用。
Objective To construct eukaryotic recombinant sTRAIL expression vector and investgate its activity against C6 glioma cell line. Methods After construction of eukaryotic recombinant sTRAIL expression vector- PcDNA-sTRAIL by means of PCR and directed cloning, the C6 glioma cell line was transfeeted by cationic liposome and the proliferation and apoptosis of cells was detected by FCM. Results The construction of eukaryotic recombinant sTRAIL expression vector- PcDNA-sTRAIL confirmed a success by gene sequencing and western blot. After transfected by cationic liposome, 50 percents of C6 cells expressed possitive protein. The apoptosis rate of C6 cell was significant higher in PcDNA-sTRAIL plus Lipofectin transfeeted group than that in three control groups （t=9.52, 7.20, 2.84 respectively, P〈0.05）. Conclutions An eukaryotic recombinant sTRAIL expression vector was successfully constructed and its anti-C6-glioma activity was confirmed in vitro study.
出处
《全科医学临床与教育》
2008年第5期380-382,F0003,F0002,共5页
Clinical Education of General Practice
基金
浙江大学医学院中青年科研启动基金资助项目(491020-542959)
关键词
胶质瘤
基因治疗
真核表达载体
转染
glioma
gene therapy
eukaryotic expression vector
transfection
