论文部分内容阅读
目的原核表达、纯化DENV-2贵州分离株NS1部分序列表达蛋白,并制备其多克隆抗体以研究其功能。方法合成DENV-2M株NS1部分基因序列,克隆到原核表达载体pET28(a),转化大肠埃希菌BL21(DE3);用IPTG诱导表达,Ni柱亲和层析纯化、回收融合蛋白;用纯化融合蛋白免疫BALB/c鼠,制备多克隆抗体,采用ELISA法检测抗体效价。结果原核表达了NS1部分序列融合蛋白,并获得了其多克隆抗体,ELISA检测抗体效价为1∶800。Western blot检测该蛋白能被His标签抗体识别。结论 DENV-2M株NS1部分序列表达蛋白可诱导小鼠产生较高效价和特异性的多克隆抗体,为进一步研究DENV-2M株NS1部分序列表达蛋白的免疫原性奠定了基础。
Objective To express and purify the partial NS1 protein of Guizhou isolates from DENV-2 and to prepare its polyclonal antibody to study its function. Methods The partial sequence of NS1 gene of DENV-2M strain was synthesized and cloned into prokaryotic expression vector pET28 (a) and transformed into Escherichia coli BL21 (DE3). The recombinant plasmid was induced with IPTG and purified by Ni affinity chromatography. The fusion protein was used to immunize BALB / c mice to prepare polyclonal antibody. The antibody titer was measured by ELISA. Results The prokaryotic expression of NS1 partial sequence fusion protein was obtained and the polyclonal antibody was obtained. The titer of the antibody was 1: 800 by ELISA. Western blot detection of this protein can be His-tag antibody recognition. Conclusion The partially expressed NS1 protein of DENV-2M strain can induce polyclonal antibody with high titer and specificity in mice, which lays the foundation for further study on the immunogenicity of NS1 protein expressed in DENV-2M strain.