组织型纤溶酶原激活剂突变体基因转染细胞株的建立

北京大学学报（自然科学版）

组织型纤溶酶原激活剂突变体基因转染细胞株的建立

李敏¹, 王玉炯¹, 崔景荣², 扈荣良², 张守峰³

¹宁夏大学生命科学学院, 银川, 750021；²北京大学医学部天然药物与仿生药物国家重点实验室, 北京, 100081；³中国人民解放军军需大学研究所，长春，130062

收稿日期:2003-03-08 出版日期:2004-07-20 发布日期:2004-07-20

Construction and Identification of t-PA Variants Gene Transfected Cell Lines

LI Min¹, WANG Yujiong¹, CUI Jingrong², HU Rongliang³, ZHANG Shoufeng³

¹The Life Science School of Ningxia University, Yinchuan, 750021; ²The State Key Laboratory of Natural and Biomimetic Drugs of Peking University Health Science Center, Beijing, 100083; ³The Institute of the Quartermaster University of PLA, Changchun, 130062

Received:2003-03-08 Online:2004-07-20 Published:2004-07-20

摘要/Abstract

摘要： 以脂质体介导法将含有t-PA突变基因的真核表达载体pCSRA及pCSRK分别转染CHO-dhfr^-，经G418及DMEM双重选择，获得两者的稳定表达细胞株。结果表明，表达产物具有良好溶纤活性；Western印迹实验证明，产物具有特异抗原性；经多次传代，细胞株具有良好的稳定性。比较了无血清培养基CHO-S-SFMⅡ与常规培养基DMEM培养条件下pCSRK细胞株产物的表达量，前者约为后者的2倍，表明无血清培养基SFM有可能用于药用性蛋白的生产。实验为t-PA新型突变体的临床应用打下一定基础。

关键词: 突变体, 稳定表达, 重组纤溶酶原激活剂

Abstract: The eucaryotic expression vector pCSRA and pCSRK, each of which included different t-PA mutant gene, were transfected into CHO-dhfr^- independently by liposome-mediated method; then through diplex choice of G418 and DMEM, the authors had got the stable cell lines of the two. Monitoring result showed that the expression products had good cellulolytic activity; the result of Western-blotting showed that they had the specific antigenicity; and passaged for many times, the cell lines had heredity stability to a certainty. For the expression quantity of the target product of pCSRK in CHO-S-SFMⅡ was as twice as in regular medium DMEM, it showed that SFM would be used to produce the medicinal protein. Thus it had done spadework under certain degree for the usages of new-type mutants of t-PA in clinic.

Key words: variants, stable expression, recombinant tissue-type plasminogen activator

中图分类号:

Q786

李敏, 王玉炯, 崔景荣, 扈荣良, 张守峰. 组织型纤溶酶原激活剂突变体基因转染细胞株的建立[J]. 北京大学学报（自然科学版）.

LI Min,WANG Yujiong,CUI Jingrong,HU Rongliang,ZHANG Shoufeng. Construction and Identification of t-PA Variants Gene Transfected Cell Lines[J]. Acta Scientiarum Naturalium Universitatis Pekinensis.

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https://xbna.pku.edu.cn/CN/Y2004/V40/I4/583

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