禽流感病毒NS2蛋白的原核表达及多克隆抗体的制备

摘要

为制备禽流感病毒(AIV) NS2蛋白的多克隆抗体,本研究将人工合成的NS2基因克隆至表达载体pET-28a中,转化大肠杆菌BL21(DE3),经IPTG诱导表达His-NS2重组蛋白.SDS-PAGE和western blot试验表明,该重组蛋白获得大量表达,可溶性高,并且具有较好的反应原性.将纯化后的重组蛋白免疫新西兰白兔制备多克隆抗体,并通过间接ELISA检测其效价达1:20 000以上.Western blot和间接免疫荧光试验显示,多克隆抗体能够与NS2蛋白特异性结合.%To prepared polyclonal antibody against NS2 protein of avian influenza virus (ATV), the NS2 gene was synthesized and cloned into pET-28a to construct the prokaryotic expression plasmid pET-NS2. The soluble rccombinant protein was expressed in E. coli BL21 (DE3) with IPTG induction and identified by the SDS-PAGE and western blot. The polyclonal antibody was prepared from the rabbit immunized with purified recombinant protein. The titer of the antibody was about 1:20 000 by detection of indirect ELISA. The western blot and indirect immunofluorescence assay also confirmed that the polyclonal antibody reacted specially with ATV NS2 protein, which would be used for study of the NS2 function.