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采用巢式PCR(nested PCR)的方法从Raji细胞中扩增出人CD80 cDNA,并将这一cDNA克隆入载体pUC19中。经PCR扩增和限制性酶切分析,进行初步鉴定后,再将人CD80 cDNA克隆到pcD-NA表达载体上,构建成为pCD-CD80重组质粒。应用脂质体介导的基因转移技术将这一表达载体导入小鼠黑色素瘤B16细胞后,用SABC法进行瞬时表达的检测。结果表明,转染后48h就具有明显人CD80基因的表达产物。
The human CD80 cDNA was amplified from Raji cells by nested PCR and this cDNA was cloned into the vector pUC19. After preliminary identification by PCR amplification and restriction enzyme digestion, the human CD80 cDNA was cloned into the pcD-NA expression vector and constructed into a pCD-CD80 recombinant plasmid. This expression vector was introduced into mouse melanoma B16 cells using a liposome-mediated gene transfer technique and transient expression was detected using the SABC method. The results showed that there was a clear expression product of human CD80 gene 48 h after transfection.