CD59-linker-CD2融合基因真核表达系统的构建  

Construction of a eukaryotic expression vector containing CD59-linker-CD2 fusion gene

作　　者：钟丹丹[1] 高美华[1] 李伟伟[1] 张蓓[1] 

机构地区：[1]青岛大学医学院免疫学教研室,青岛医学硕士研究生266071

出　　处：《医学研究生学报》2011年第1期15-19,共5页Journal of Medical Postgraduates

基　　金：国家自然科学基金(30972677)

摘　　要：目的 CD59和CD2都是重要的淋巴细胞膜表面分子,参与T细胞信号转导。文中构建CD59和CD2融合基因真核表达系统,探讨CD59向T细胞传递信号的机制及CD59与CD2的相互作用。方法在CD59基因和CD2基因间加入一段柔性链接头(linker)基因序列,构建CD59-linker-CD2融合基因真核表达系统。利用RT-PCR法从人T细胞白血病细胞系Jurkat细胞总RNA中扩增CD59和CD2基因,分别构建CD59-T和CD2-T重组克隆载体,经限制性核酸内切酶酶切后与pIRES质粒进行重组,构建pIRES-CD59-linker-CD2融合基因真核表达系统。结果测序证实CD59和CD2基因扩增成功;经限制性核酸内切酶酶切、PCR和测序鉴定表明携带CD59-linker-CD2融合基因的重组质粒构建成功。结论构建CD59-linker-CD2真核表达系统为在真核细胞中表达CD59与CD2并进一步探讨其在T细胞信号转导中的作用提供了实验材料。Objective Both CD59 and CD2 are important molecules on the surface of lymphocytes and involved in T cell signal transduction.The purpose of this study was to construct a eukaryotic expression vector of the CD59 and CD2 fusion gene to facilitate the investigation of the mechanism of signal transduction from CD59 to T cells and the interaction between CD59 and CD2.Methods We constructed a eukaryotic expression vector of the CD59-linker-CD2 fusion gene by linking the CD59 and CD2 genes with a flexible chain（linker）.The CD59 and CD2 genes were cloned by RT-PCR from the total RNA of Jurkat T cells to establish CD59-T and CD2-T cloning vectors,respectively,which in turn were cloned into the plasmid pIRES following restriction endonuclease digestion to construct a eukaryotic vector containing the CD59-linker-CD2 fusion gene.Results DNA sequencing confirmed the successful amplification of the CD59 and CD2 genes.Restriction endonuclease digestion,PCR and DNA sequencing analysis proved that the eukaryotic expression vector containing the CD59-linker-CD2 fusion gene was constructed correctly.Conclusion The successful construction of the eukaryotic expression vector containing the CD59-linker-CD2 fusion gene has prepared the experimental ground for further study of the expressions of CD59 and CD2 in eukaryotic cells and their role in T cell signal transduction.

关 键 词：CD59基因 CD2基因 融合基因 真核表达载体 

分 类 号：R392.12[医药卫生—免疫学]