| HuCTLA4Ig真核表达载体的构建及其表达 |
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| 引用本文： | 祁小平,钱军华,谢晋,马宝骊. HuCTLA4Ig真核表达载体的构建及其表达[J]. 现代免疫学, 2000, 20(5): 282-285 |
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| 作者姓名： | 祁小平 钱军华 谢晋 马宝骊 |
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| 作者单位： | 1. 上海第二医科大学,上海市免疫学研究所,上海,200025
2. 江苏金丝利药业有限公司,宜兴,214205 |
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| 基金项目： | 国家自然科学基金资助项目(基金编号39770705) |
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| 摘 要： | 用RT PCR方法从人外周血单个核细胞总RNA中分别扩增出CTLA4的胞膜外区基因和Igγ1恒定区基因 ,将它们拼接后插入质粒pcDNA3。将重组的HuCTLA4Ig/pcDNA质粒转染CHO细胞 ,经筛选得到了高效、稳定表达HuCTLA4Ig的克隆。培养上清经ProteinA亲和层析柱纯化后 ,所得蛋白在非还原条件下SDS PAGE呈分子量为 11kD的一条带 ;FACS检测显示该纯化蛋白与B7分子有良好的结合能力 ;在体外能抑制MLR的增殖反应。
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| 关 键 词： | HuCTLA4Ig 真核表达 CHO细胞 |
| 文章编号： | 1001-2478(2000)05-0282-04 |
| 修稿时间： | 1999-12-06 |
Construction of Eukaryotic Expression Vector Containing HuCTLA4Ig and its Expression in Mammalian Cells |
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| Qi Xiaoping,Qian Junhua,Xie Jin,Ma Baoli. Construction of Eukaryotic Expression Vector Containing HuCTLA4Ig and its Expression in Mammalian Cells[J]. Current Immunology, 2000, 20(5): 282-285 |
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| Authors: | Qi Xiaoping Qian Junhua Xie Jin Ma Baoli |
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| Affiliation: | Qi Xiaoping;(Shanghai Institute of Immunology, Shanghai Second Medical University, Shanghai 200025);Qian Junhua;(Jiangsu Kingsley Pharmaceutical Co. LTD, Yixin 214205);Xie Jin,Ma Baoli;(Shanghai Institute of Immunology, Shanghai Second Medical University, Shanghai 200025) |
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| Abstract: | The extracellular region of the human CTLA4 cDNA and the constant region of the human Igγ1 cDNA were amplified by RT-PCR,and the amplified cDNAs were linked and inserted into pcDNA3 plasmid The plasmid HuCTLA4Ig/pcDNA was transfected into CHO cells After series of screening procedure,several transfected CHO clones were obtained,which expressed HuCTLA4Ig in high level and constitutively The culture suspension was purified with affinity chromatography using protein A column The result of nonreducing SDS-PAGE demonstrated that the recombinant fusion protein appeared a signal band of 11kD The FACS analysis and inhibition assay of MLR showed that the protein could authentically bind to B7 molecule |
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| Keywords: | HuCTLA4Ig eukaryotic expression CHO cell |
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