融合表达IL-2和EGFP逆转录病毒载体的构建

Construction of Retrovirus Vector for Fusion Expression of IL-2 and EGFP

目的构建融合表达IL-2和EGFP报告基因的逆转录病毒载体。方法设计载体pRevTet-On和pEGFP-C1的接头序列,经酶切连接重组为过渡载体pRevEGFP-C1,同时对质粒pBV220-IL-2和pEGFP-C1进行酶切,连接重组为过渡载体pEGFP-C1-IL-2。分别酶切质粒pRevEGFP-C1和pEGFP-C1-IL-2,琼脂糖凝胶电泳回收目的片段IL-2,并将其连接到pRevEGFP-C1的多克隆位点(MCS)中,转化至E.coliDH5α进行扩增,提取质粒DNA获得重组体pRevEGFP-C1-IL-2。结果经分析鉴定,所构建的重组载体pRevEGFP-C1-IL-2完全正确。结论已成功地构建了融合表达IL-2和EGFP的逆转录病毒载体pRevEG-FP-C1-IL-2。

Objective To construct a retrovirus vector for the fusion expression of IL-2 and EGFP. Methods Design the adaptor of plasmids pRevTet-On and pEGFP-Cl ,digest with restriction endonuclease and ligate to recombine a transition retrovirus vector pRevEGFP-Cl. Meanwhile,digest plasmids pBV220-IL-2 and pEGFP-Cl with restriction endonuclease and ligate to recombine another transition retrovints vector pEGFP-Cl-IL-2. Digest plasmids pRevEGFP-Cl and pEGFP-Cl-IL-2 with restriction endonuclease respectively. Recover target gene fragment IL-2 by agarose gel electrophoresis,insert into vector pRevEGFP-Cl and transform to E. coil DH5α for proliferation. Extract plasmid DNA to obtain a recombinant pRevEGFP-Cl-IL-2. Results Analaysis proved that vector pRevEGFP- Cl-IL-2 was correctly constructed. Conclusion A retrovirus vector pRevEGFP-Cl-IL-2 for the fusion expression of IL-2 and EGFP was successfully constructed.