摘要
为成功表达猪繁殖与综合征病毒(PRRSV)结构蛋白GP5,通过PCR方法从重组质粒pGEM ORF5扩增得到缺失N端疏水序列的基因片段dORF5(deletingORF5)。将dORF5克隆至原核高效表达载体pGEX 4T 2,在E.coliBL21细胞中成功表达了重组蛋白GST dORF5,表达产物以包涵体的形式存在,表达量为20 8%。Western Blot结果表明重组蛋白可被PRRSV阳性血清所识别。利用融合肽进行亲和层析得到高纯度的重组蛋白,为进一步研究PRRS病毒结构蛋白的结构和功能奠定了基础。
In order to successfully express the structural protein GP5 of porcine reproductive and respiratory syndrome virus(PRRSV),the gene segment dORF5 deleting N-terminal very hydrophobic sequence were successfully amplified from recombinant plasmid pGEM-ORF5 by PCR and were cloned into prokaryotic expression vector pGEX-4T-2.The recombinant fusion proteins GST-dORF5 were highly expressed in E coli.cell BL21 in the forms of inclusion bodies and could amount to 20.8% of the total mass of bacterial proteins.Western-Blot showed the recombinant protein could react with the porcine polyclonal antibodies against PRRSV.Recombinant proteins were purified and renatured.The purity of recombinant proteins affinity-purified using Glutathione Sepharose 4B were above 90%.The studies provided fundamental data and materials for the further study on the structure and function of structural proteins of PRRSV.
出处
《畜牧兽医学报》
CAS
CSCD
北大核心
2004年第1期64-69,共6页
ACTA VETERINARIA ET ZOOTECHNICA SINICA
基金
霍英东青年教师基金(71032)
