蓝舌病病毒VP7蛋白的原核表达及免疫原性鉴定

Prokaryotic Expression and Immunogenicity Identification of Bluetongue Virus VP7 Protein

为获得蓝舌病病毒(bluetongue virus,BTV)25型的VP7原核表达蛋白,本试验以蓝舌病病毒重组质粒为模板,PCR扩增VP7基因,将其克隆于pET-24b(+)表达载体中,获得pET-24b-BTV-VP7重组质粒。经酶切和测序鉴定后,转化大肠杆菌BL21(DE3)受体菌,IPTG诱导表达His-BTV-VP7蛋白。在变性条件下用镍亲和层析柱纯化His-BTV-VP7蛋白,经Western blotting及ELISA鉴定其免疫原性。结果显示,His-BTV-VP7蛋白以包涵体形式表达,大小约为40ku;Western blotting和ELISA检测此原核表达蛋白能与山羊阳性血清发生特异性反应,具有良好的免疫原性。本研究为后续建立蛋白芯片检测方法奠定了基础。

To obtain VP7 protein of bluetongue virus （25 type）,VP7 gene was amplified and cloned in pET-24b （＋） expression vector. The pET-24b-BTV-VP7 recombinant plasmid was transformed into BL21 （DE3）, then the VP7 protein of bluetongue virus was expressed using IPTG and purified by nickel affinity chromatography in vitro. Immunogenicity of VP7 protein was determined by Western blotting and ELISA. The results showed that the molecular weight of VP7 protein was about 40 ku and it could react with goat positive serum specifically. This study laid the foundation for establishing protein chip detection methods in the future.