rhIFN-β1a cDNA在CHO细胞上的表达

Expression of Recombinant Human IFN-β1a cDNA in CHO Cells

目的 在CHO细胞上表达rhIFN βcDNA。 方法 利用脂质体技术 ,将含hIFN β基因的pRC CMV IFNβ DNA载体和pSV2dhfr DNA质粒按 3∶1的比例转染CHO k1dhfr 细胞 ,后者是选择性标记。筛选在缺乏胸腺嘧啶的选择培养基生长的单个细胞克隆 ,通过MTX加压扩增与dhfr基因共转染IFNβ基因。筛选的细胞株通过大规模培养 ,收集培养液经一系列的步骤纯化。结果 能够耐受 0 .6 μmol LMTX的细胞可产生IFN 10 5unitperday 10 6cells ,CHO细胞中表达的hIFN β纯化后的抗病毒比活性可达 2 .2× 10 8IU mg。结论 建立了适合表达人干扰素

Objective To express recombinant human IFN-β1a cDNA in CHO cells.Methods Mix recombinant plasmids pRC/CMV-IFNβ DNA and pSV2dhfr-DNA (used as a selective marker) at a ratio of (3∶1) and transfect into CHO-kldhfr cells by liposome technique. Screen the single clones grown in thymine-free selective medium for the pressure amplification of IFN-β gene. Subject the screened clones to large-scale culture, and collect the culture liquid for the purification by a series of procedures.Results The daily yield of IFN-β secreted by the cells resistant to 0.6 μmol/L MTX was 10~5 unit/10~6 cells.The specific activity of human IFN-β expressed in CHO cells reached 2.2×10~8 IU/mg after purification.Conclusion A CHO cell strain suitable for the expression of human IFN-β was established.