HLA—A＊0201的cDNA克隆及在COS—7细胞上的瞬时表达

Cloning of HLA-A * 0201 and its transient expression on COS-7 cells

目的 :克隆HLA A 0 2 0 1编码区的cDNA ,并建立在COS 7细胞上的瞬时表达系统。方法 :采用逆转录 PCR技术从HLA A 0 2 0 1阳性的淋巴细胞中获得HLA A的cDNA ,将其定向克隆至载体pBluescriptIISK(+ / - )中 ,经测序确证后 ,将该基因插入真核表达载体pcDNA3中 ,通过阳离子脂质体介导转染至COS 7细胞 ,用流式细胞仪测定细胞表面HLA A 0 2 0 1分子的瞬时表达。结果 :获得的HLA A 0 2 0 1基因的cDNA序列与Genebank登录的cDNA序列一致。FACS检测结果显示 ,COS 7细胞上HLA A 0 2 0 1的表达率为 5 3%。结论 :成功克隆HLA A 0 2 0 1基因 ,并实现了在COS 7细胞上的瞬时表达 ,为研究HLA

Objective:To clone HLA A *0201 cDNA and express it transiently on COS 7 cells.Methods:HLA A cDNA isolated from the HLA A *0201 positive lymphocytes by using RT PCR was cloned into pBluescript II SK vector directionally.After the sequence was confirmed,the cDNA was inserted into mammalian expression vector pcDNA3.The recombinant vector was transfected into COS 7 cells by cation liposome.The transient expression on the cells was measured by flow cyteometer.Results:The cDNA was identical with HLA A *0201 cDNA published on Genebank.Determined by flow cytemeter,the expressing rate was recorded for 57%.Conclusion:We have cloned HLA A *0201 successfully and expressed it transiently on COS 7 cell,which would be potentially useful in research on killing tumor restricted by HLA A2.