人促红细胞生成素在CHO细胞上的表达  

Expression of Erythropoietin in CHO Cells

作　　者：梅柱中[1] 岳军明[1] 张茂林 乔贵林 殷震[1] 

机构地区：[1]长春农牧大学基因工程实验室,长春130062

出　　处：《药物生物技术》2000年第3期129-131,共3页Pharmaceutical Biotechnology

基　　金：全军医药卫生重点项目! ( 96Z0 5 0 )

摘　　要：为了获得人促红细胞生成素 (EPO)的高效表达 ,以EPO gDNA为基础构建了两个真核表达载体pDE、pCE ,将它们分别经脂质体转染CHO dhfr-细胞 ,其 48h瞬时表达量分别为 2 7 5ng/ml与 88 90 ng/ml。然后用氨甲喋呤 (MTX)对转染细胞进行加压并挑选细胞克隆 ,共获得 3株高效表达EPO的工程细胞株A ,B ,C ,其中A来自质粒 pCE ,B和C来自 pDE ,在 2× 10 -6mol/L的MTX的压力下 ,它们 48h的最高表达水平分别为A :8 7μg/ml,B :10 76 μg/ml,C :16 44 μg/ml。To achieve high expression of erythropoietin,two eukaryotic vectors pDE and pCE were constructed with EPO gDNA. They were transfected into Chinese Hamster Ovary cells by lipofectamine respectively. Their transient expression in 48 h were 27 5 ng/ml and 88 9 ng/ml. The transfected cells were selected by methotrexate. With the increase of MTX concentration step by step,the exprssion level of EPO were also increasing. After about 2 months,three stable cell lines (one from pCE,the other two from pDE)were obtained. Under the pressure of 2×10 6 mol/L MTX,their secretion in 2 days were up to 8 70 μg/ml,16 44 μg/ml and 12 65 μg/ml, respectively. By reticulacyte counting,these secreted EPO all had biological activity in vivo .

关 键 词：人促红细胞生成素 真核表达 CHO细胞 抗贫血药 

分 类 号：R973.3[医药卫生—药品] Q78[医药卫生—药学]