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为了获得能用于抗虹鳟(Rainbow trout,学名,Oncorhynchus mykiss)病毒性疾病生物制剂开发的高纯度Ⅰ型干扰素(rtIFN3)蛋白,克隆了虹鳟rtIFN3,构建了pET21a-rtIFN3/BL21(DE3)原核表达与发酵系,并对rtIFN3进行了表达。重组的rtIFN3分子质量约18.5ku,以包涵体形式表达,表达量占全菌蛋白量的16%。随后,提取2L发酵瓶的包涵体蛋白,经变性与复性后,再经分子筛和阴离子交换层析进行纯化。纯化后,rtIFN3SDS-PAGE电泳结果表明蛋白纯度达98%,复性率为3%。本研究获得的高纯度复性蛋白为生物制剂的研制提供了材料。
In order to obtain rtIFN3 protein, which can be used in biological preparations of rainbow trout (Oncorhynchus mykiss), the rainbow trout rtIFN3 was cloned and the prokaryotic expression vector pET21a-rtIFN3 / BL21 (DE3) was constructed Expression and fermentation system, and rtIFN3 expression. Recombinant rtIFN3 molecular weight of about 18.5ku, expressed in the form of inclusion bodies, the expression amount of 16% of the total amount of protein. Subsequently, the inclusion protein of 2 L fermentation flask was extracted, denatured and refolded, and then purified by molecular sieve and anion exchange chromatography. After purification, rtIFN3SDS-PAGE electrophoresis results show that the protein purity of 98%, the refolding rate of 3%. The high-purity renatured proteins obtained in this study provide materials for the development of biological agents.