人红细胞生成素工程细胞株的特性鉴定

The Analysis and Identify of Characteristics for the Engineering Cell Lines of Erythropoietin

目的 :为了对人红细胞生成素工程细胞株 (F10 - 6 )进行特性鉴定。方法 :将人红细胞生成素基因构建的重组表达质粒pCDB与标志质粒pSV -dhfr共转染CHO -dhfr- 细胞中 ,经氨甲喋呤加压筛选和亚克隆挑选 ,获得高效稳定表达EPO的工程细胞株并对其进行鉴定。结果 :实验结果表明 ,工程细胞株无细菌、支原体、真菌和病毒污染 ;并无致瘤性 ;染色体数目不变 ;连续传代5 0次和三次冻存复苏后 ,细胞表达稳定。结论 :该工程细胞株可用于工业化生产。

Purpose:The aim was to identify the characteristic of engineering cell of human erythropoietin.Methods:The recombinant plasmid pCDB constructed with EPOcDNA and Marked plasmid pSV-dhfr was cotransfected into CHO-dhfr^- cell.The cloneof the highest expression EPO were selected and identified through methotrexate amplification and subclone.Results:The results of identification show that F10-6 cell line have not been contaminated by mycoplasma,bacteria,fungi,and virus.It also have no the form of tumor in nude mice.The chromosomal number of F10-6 cell line is normal.It was bequeathed for fifty passages and resuscitated after freezing three times.The expression level of EPO is stable.Conclusion:F10-6 engineering cell can be used for the production of industrialization.