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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。经网织红细胞法测得它们均具有体内生物活性
In order to obtain the high expression of human erythropoietin (EPO), two eukaryotic expression vectors, pDE and pCE, were constructed on the basis of EPO gDNA, and were transfected into CHO dhfr- cells by liposome respectively. The 48h transient expression Respectively, 275 ng / ml and 88 90 ng / ml. Then, the transfected cells were treated with methotrexate (MTX) and the cell clones were selected. A total of 3 highly expressed EPO-expressing cell lines A, B and C were obtained, in which A was from plasmid pCE, B and C were from pDE, Under the pressure of 10 -6 mol / L MTX, the highest expression levels of them at 48 h were A: 8 7 μg / ml, B: 10 76 μg / ml and C: 16 44 μg / ml, respectively. They all have in vivo biological activity as measured by reticulocyte method