【摘要】 采用基因克隆与腺相关病毒包装技术,构建人Th1细胞特异性转录因子T-bet基因的腺相关病毒载体,并转染入胃癌细胞系SGC-7901细胞。检测转染细胞IFN-γ的分泌功能及其对SGC-7901细胞生物学作用的影响。结果:(1)获得含有T-bet基因的重组腺相关病毒(rAAV-eGFP-T-bet);(2)用rAAV-eGFP-T-bet感染SGC-7901细胞后,约30%~40%的细胞出现绿色荧光。对被感染细胞进行RT-PCR,扩增出1 670 bp的T-bet,并经Westernblot证实;(3)转染后的细胞测得388 bp的IFN-γ目的条带和IFN-γ蛋白的表达。由此表明,人T-bet基因的腺相关病毒载体rAAV-eGFP-T-bet,可成功转染SGC-7901,并致使转染细胞分泌IFN-γ。为进一步开展T-bet基因干预治疗肿瘤的研究奠定了基础。更多还原

【Abstract】 In order to investigate the effect of T-bet on malignant cells,we selected SGC-7901,a kind of human gastric carcinoma cell line,and used gene clone technique and adeno-associated virus(AAV) packing technology,thus obtaining a recombinant rAAV-eGFP-T-bet and T-bet gene-transfected SGC-7901 cells.Then the function of T-bet gene-infected SGC-7901 cells was researched by detecting the levels of IFN-γ and T-bet production.The results showed:(1)It was verified that rAAV-T-bet’s packing was completed;(2)After SGC-7901 cells was transfected by rAAV-eGFP-T-bet,a green fluorescence was found in about 30%-40% SGC-7901s,and the gene of 1 670 bp(T-bet) and 388 bp(IFN-γ) were generated from SGC-7901s cells;(3) The proteins of IFN-γ and T-bet secreted by SGC-7901 cells were also detected.These reveal that SGC-7901 cell is efficiently infected by rAAV encoding T-bet,which can induce transfected cells to secret IFN-γ.It may be useful in the researches on cancer immune therapy of transfecting T-bet gene.更多还原