稳定表达结核分枝杆菌38抗原的EL-4细胞株的建立及鉴定

Establishment of EL-4 cell line stably expressing 38 antigens of Mycobacterium tuberculosis

将结核分枝杆菌的38抗原基因片段经PCR方法扩增并插入到pcDNA3真核表达载体中,通过脂质体转染EL-4细胞,经G418筛选,用RT-PCR方法和ELISA方法检测整合和表达情况。结果成功地构建了pcDNA3-38抗原重组质粒,转染细胞中可检测到38抗原的存在,PCR扩增也证实38抗原基因已稳定整合于EL-4细胞的染色体中。本实验为今后在小鼠体内检测结核DNA疫苗激发的细胞毒性T淋巴细胞(CTL)反应奠定了基础。

The 38 antigen cDNA of Mycobacterium tuberculosis was amplified and inserted into pcDNAS eukaryotic expressing vector. The recombinant plasmid was transfected into EL-4 cell line by liposome and screened by G418. The intergra-tion and expression of 38 antigen cDNA was detected by RT-PCR and ELISA respectively. The results showed that recombinant plasmid pcDNA3-38 was successfully constructed. It further confirmed that 38 antigen cDNA had been stably integrated into the chromosome of EL-4 cell by RT-PCR method,and that 38 antigen could express stably in transfected EL-4 cells. This study provided basis for detecting the cytotoxic T-lymphocyte (CTL) reaction stimulated by DNA vaccine of tuberculosis in rats.