重组猪干扰素α的纯化工艺研究  

Purification of Recombinant Porcine Interferon α

作　　者：苏世云[1] 夏俊保[1] 赵俊[1] 吴琼[1] 王明丽[1] 

机构地区：[1]安徽医科大学,安徽合肥230032

出　　处：《安徽农业科学》2010年第14期7363-7365,共3页Journal of Anhui Agricultural Sciences

基　　金：2008年度安徽省教育厅自然科学基金重点项目(KJ2008-A085)

摘　　要：[目的]研究重组猪干扰素α的纯化工艺,为进一步研究该蛋白的分子结构及rPoIFN-α标准品的制备奠定基础。[方法]将含重组菌E.coliBL21诱导表达后得到的粗制rPoIFN-α,分别用2种方案进行纯化。方案1,依次用GST亲和层析法、DEAE阴离子交换法及分子筛法层析三步法进行纯化;方案2,依次用GST亲和层析和分子筛法层析两步法进行纯化。纯化结果用SDS-PAGE及Western-blot进行纯度检测及鉴定。[结果]诱导表达后的表达产物经SDS-PAGE检测,发现在45Kd分子量处有目的条带,Western-blot检测有特异性条带。通过两步法纯化后的rPoIFN-α纯度达96%,蛋白得率为42.8%,三步纯化纯度为98.8%。[结论]两步法得到的蛋白纯度非常接近三步法,但与三步法相比工艺更为简单方便。[Objective]To Study the purification of recombinant porcine interferon α（rPoIFN-α）,establish a foundation of study on the structure of the rPoIFN-α and production of standard preparation of the protein. [Methods]The rPoIFN-α was induced and extracted from the recombinant E. coli BL21,and then purified by two strategies. The first strategy was that the rPoIFN-α was purified by GST affinity chromatography,ion exchange chromatography and gel filtration in turn. The second strategy was that the rPoIFN-α was purified by GST affinity chromatography and gel filtration in turen. The purification was detected by SDS-PAGE,and rPolFN-α was identified by Western-blot. [Result]After the expression of inducible expreesion,the target strip was detected by SDS-PAGX in 45 kDa,and the specific strip was detected using Western-blot. The purification of purified rPoIFN-α get from the two-step method and the three-step method. ware about 96% and 98. 8% ,respectively. The protein extractien yield was 42. 8%. [Conclusion]The purified rPoIFN-α get from the first strategy was about 96% of its purification,this strategy is more convenient than the second strategy,and more fit for semi-works production.

关 键 词：重组蛋白 猪Α干扰素 层析 

分 类 号：S855.3[农业科学—临床兽医学]