人β干扰素基因定点突变及在sf9细胞中的表达和活性研究  

Mutation and Expression of Human IFNβ in sf9 Cells and Study of Its Bioactivity

作　　者：李晓阳[1] 郭学青[2] 

机构地区：[1]首都医科大学附属北京朝阳医院京西院区检验科,100043 [2]北京军区总医院263临床部检验科,101149

出　　处：《国际检验医学杂志》2010年第10期1096-1098,共3页International Journal of Laboratory Medicine

摘　　要：目的研究人β干扰素突变体在杆状表达系统中的高效表达,为新型基因药物的研制提供可行性。方法采用PCR定点突变技术构建人干扰素bS17突变基因,克隆到pFastBacTMHTB上,获得重组转移载体pFastBac-IFNβS17。将pFast-Bac-IFNβS17转化入DH10BacTM细胞,得到Bacmid-IFNβS17。将Bacmid-IFNβS17转染sf9细胞,筛选到重组病毒Bac-IFNβS17。将Bac-IFNβS17感染sf9细胞进行表达。结果 Southern-blot证明IFNβS17已插入重组病毒中(3.9kb左右的杂交带);SDS-PAGE、蛋白质印迹证明IFNβS17在sf9细胞中得到了表达。实验表明sf9细胞中120h的表达产物干扰素活性最高,为2.62×105IU/mL。结论突变基因在sf9细胞得到了高效表达,且表达的重组蛋白具有IFN的生物活性。Objective The expressed recombinant protein have full bioactivity of hlFNβto investigate the feasibility of research and development a new gene drug. Methods The baculovirus shuttle vector,pFastBac-IFNβS17 was constructed which contains the mutant gene of IFNβS17. The recombinant vector was transformed into DH10Bac^TM cells to get Bacmid-IFNβS17. The recombinant virus Bac-IFNβS17 was screened by transfecting Bacmid-IFNβS17 into sf9 cells. The sf9 cells were infected with the recombinant virus. Results The test showed that protein was successfully expressed in sf9 cells and its best bioactivities were 2.62×105IU/ml testified in 120 hours. Conclusion All the data indicated the expressed recombinant protein have full bioaetivity of hIFNβ.

关 键 词：干扰素类 细胞 基因表达 突变 

分 类 号：Q78[生物学—分子生物学]