四川大学学报（医学版）2011，Vol.42Issue(3)：317-321,5.
人血清白蛋白与促红细胞生成素融合蛋白的构建及表达
Construction and Expression of Recombinant Human Serum Albumin-EPO Fusion Protein
黄迎春 1苟兴华 2韩蕾 2李德华 2赵兰英 2吴洽庆2
作者信息
- 1. 四川大学华西医院,移植免疫重点实验室,成都,610041
- 2. 成都地奥制药集团有限公司,成都,610041
- 折叠
摘要
Abstract
Objective To construct the recombinant plasmid pCI-HLE encoding human serum album-EPO (HSA-EPO) fusion protein and to express it in CHO cell.Methods The cDNA encoding human serum album and EPO were amplified by PCR, and then spliced with the synsitic DNA fragment encoding GS (GGGGS)3 by overlap PCR extension to form LEPO.After BamH Ⅰ digestion, the HSA and LEPO was ligated to generate the fusion HSA-EPO gene and was then cloned into the expression vector pCI-neo to generate the recombinant plasmid pCI-HLE.The plasmid pCI-HLE was transfected into CHO cell by liposome protocol.Then, the recombinant cells were screened by G418 and identified by PCR and Western blot.Expression of fusion protein was evaluated by Enzyme Linked Immunosorbent Assay (ELISA).Results Restrictive enzymes digestion and DNA sequencing revealed that HSA-EPO fusion gene was cloned into expression vector pCI-neo successfully.PCR and Western blot analysis confirmed that the fusion gene was integrated in the genome of CHO cells and expressed successfully.The HSA-EPO production varied from 86 IU/(mL·106·72 h) to 637 IU/(mL · 106 · 72 h).Conclusion The results confirmed that HSA-EPO fusion gene can be expressed in the CHO cells, with EPO immunogenicity, which could serve as foundation for the development of long-lasting recombinant HSA-EPO protein.关键词
血清白蛋白/促红细胞生成素/融合蛋白/CHO细胞Key words
Human serum album/ Erythropoietin/ Fusion protein/ CHO cell引用本文复制引用
黄迎春,苟兴华,韩蕾,李德华,赵兰英,吴洽庆..人血清白蛋白与促红细胞生成素融合蛋白的构建及表达[J].四川大学学报（医学版）,2011,42(3):317-321,5.
