鹅坦布苏病毒非结构蛋白NS1的原核表达及纯化

摘要

根据鹅坦布苏病毒JS804株非结构蛋白NS1基因序列,设计1对特异性引物,利用PCR方法扩增得到完整的NS1基因,并将其克隆至原核表达载体pET28a和pET32a上,构建出重组表达质粒pET28a-NS1和pET32a-NS1,转化至BL21(DE3)中,经IPTG诱导得到NS1融合蛋白(His-NS1),其分子质量约分别为44,58 kDa.均在诱导后6h达到表达量高峰.分析显示,2种融合蛋白均以包涵体形式存在,包涵体经过变性和复性后均可获得单一、高表达量的目的蛋白,为进一步开展关于鹅坦布苏病毒NS1蛋白的研究奠定了基础.%The goose tembusu virus JS804 strain non-structural protein NS1 gene was amplified by PCR with the specific primers designed on the basis of NS1 gene and then was inserted into the prokaryotic vector pET28a and pET32a for the construction of recombinant expression plasmid pET28a-NSl and pET32a-NSl. Then pET28a-NSl and pET32a-NSl were transformed respectively into Escherkhia coli BL21 ( DE3 ). The recombinant protein NSls ( His-NSl ) were obtained with the induction of IPTG and the molecular weight of the fusion protein was 44 kDa and 58 kDa respectively. After 6 h induction by IPTG, the yield of fusion proteins reached peaks. The analysis showed that two fusion proteins were expressed in Escherichia coli into inclusion bodies. The fusion protein was purified to pure and high-level proteins by extracting the inclusion bodies with urea. The results laid the foundation for the further studies on the NS1 protein of goose tembusu virus.