CD80-IgG1Fc段融合蛋白真核表达载体的构建及表达

Construction of eukaryotic expression vector of CD80-IgG1 Fc fragment fusion protein and its expression in CHO cells

目的构建人CD80-IgG1Fc段融合蛋白的真核表达载体CD80-IgG1Fc/pcDNA3.1(+),并在CHO细胞中进行表达。方法应用T-A克隆技术和亚克隆技术,构建人CD80膜外段-IgG1Fc段融合蛋白的真核表达载体CD80-IgG1Fc/pcDNA3.1(+),并将其转染入CHO细胞株并筛选,进行稳定表达。通过Western blot及ELISA法,检测融合蛋白的表达。结果成功地构建了真核表达载体CD80-IgG1Fc/pcD-NA3.1(+),并建立CHO细胞稳定表达株。Western blot及ELISA法检测到细胞培养上清中有融合蛋白的表达。结论成功地构建了人CD80膜外段-IgG1Fc段融合蛋白的真核表达载体CD80-IgG1Fc/pcDNA3.1(+),并在CHO细胞中稳定表达,为下一步抗肿瘤的研究奠定了基础。

AIM ： To construct an eukaryotic expression vector of CD80-1gGI Fc, and to express the fusion protein in CHO cells. METHODS： The gene encoding the CD80-1gGI Fc fusion protein were constructed in eukaryotic expression vector pcDNA3. 1 （ ＋ ） by means of T-A cloning and subcloning techniques, then was transfected into CHO cells for stable expression. The expression of the fusion protein was detected by Western blot and ELISA. RESULTS： DNA sequencing and restriction endonuclease digestion analysis indicated that the eukaryotic expression vector CD80-1gGI Fc/ pcDNA3. 1 （ ＋ ）was successfully constructed. After the recombinant plasmid was transfected into CHO cells, the stable expression of the fusion protein was demonstrated by Western blot and ELISA. CONCLUSION： The eukaryotic expression vector of CD80-1gGI Fc/pcDNA3.1 （ ＋ ） was successfully constructed and stably expressed in CHO cells, providing basis anti-tumor study.