甲型流感病毒核蛋白基因的克隆表达及纯化

Clone expression and purification of influenza A virus nucleoprotein gene

目的 将甲型流感病毒核蛋白(NP)基因克隆到原核表达载体进行可溶性融合表达,制备纯化的病毒核蛋白,为制备甲型流感病毒单克隆抗体提供材料。方法 提取甲型流感病毒RNA ,设计引物,RT PCR扩增NP基因,利用基因工程的手段,将甲型流感病毒的NP基因在大肠埃希菌中进行融合表达,并将表达产物进行亲和层析。结果 成功构建了甲型流感病毒NP基因原核表达载体,经亲和层析制备了较高纯度的目的表达产物。结论 通过合理控制发酵时间、生长温度和诱导物浓度,制备了较为理想的可溶性甲型流感病毒核蛋白。

Objective To clone the influenza A virus NP gene into expression vector and to purify the target protein, which was used to study the preparation of monoclonal antibody. Methods The RNA of influenza A virus was extracted and primers were designed according the NP gene sequence, then the NP gene of influenza A virus was expressed in E.coli DH5α and the NP protein was purified by affinity chromatography. Results The recombinant expression vector-pGEX-4T-2-NP was successfully constructed and relatively pure target protein was obtained. Conclusion Through reasonably controlling the fermentation time, growth temperature and induction concentration, satisfactory soluble target product was obtained.