摘要
先将鸡贫血病毒VP3基因插入真核表达载体pVAX1的多克隆位点,再将pIRES1载体上的IRES启动子及新霉素基因一同切下插入pVAX1的VP3基因下游,然后切下新霉素基因,将hIL-18基因插入其中,构建成pVVP3IL-18,将pVVP3IL-18转染Hela细胞。间接免疫荧光表明,在细胞膜和细胞浆观察到了特异的黄绿色荧光,证明表达了hIL-18。pVVP3IL-18转染人结肠癌细胞HCT-116后,经AO/EB染色及电镜观察,可见大量细胞凋亡。说明真核重组质粒pVVP3IL-18可在HeLa细胞中表达,并可在体外诱导人结肠癌细胞HCT-116凋亡。
IRES promotor and neomycin gene from pIRES1 vector were inserted into multiple clone site of pVAX1 vector, then a recombinant plasmid pWP3IL-18 was constructed by inserting CAV VP3 gene and human IL-18 gene into multiple clone site and neomycin site , respectively. The recombinant plasmid pVVP3IL-18 was transfected to Hela cells and the results of indirect immunofluorescence showed that specific bright yellow green flurescene was observed around the membrane and across the plasma of the transfected Hela cells when tested with anti-mouse Ig G, FITC conjugate. The results of AO/EB staining and electron microscope showed apoptosis was induced greatly after pVVP3IL-18 was transfected to HCT-116 cells. The above results showed that the recombinant plasmid pVVP3IL-18 can express in Hela cells and can induce apoptosis in HCT-116 cells.
出处
《高技术通讯》
EI
CAS
CSCD
2003年第9期43-46,共4页
Chinese High Technology Letters
基金
973规划(G1999011902)资助项目。
