马传贫驴白细胞弱毒疫苗株酸性蛋白p9基因的克隆与表达

从感染驴白细胞的马传贫驴白细胞弱毒疫苗株前病毒DNA中克隆了编码p9蛋白的基因,并在大肠杆菌中进行了表达。所表达的蛋白是一种可溶性的融合蛋白,其氨基端带有6个组氨酸的标签,因此可以用固定化金属离子亲和层析法在非变性条件下进行纯化。在间接酶联免疫吸附试验(ELISA)和免疫印迹试验中,重组的酸性蛋白p9可与马传贫阳性血清样品发生反应,而与健康马血清无任何反应。这表明该重组蛋白具有良好的抗原性和特异性,可用于马传贫弱毒疫苗株在体内外复制及在接种马体内免疫应答的研究 。

Cloning and Expression of Acidic Protein p9 from Donkey Leukocyte-attenuated Vaccine Strain of Equine Infectious Anemia Virus

The gene coding for the acidic protein p9 of the donkey leukocyte-attenuated vaccine strain of equine infectious anemia virus (EIAV) was cloned from its proviral DNA obtained from the vaccine strain infected donkey leukocyte cultures,and expressed in E.coli.The protein was expressed in a soluble form and as a fusion protein.There was a 6 histidines tag at its amino acid terminus.It was purified by immobilized metal ion affinity chromatography under native conditions.Recombinant acidic protein showed reactivity to EIAV positive serum samples and no reactivity to normal horse sera in indirect enzyme-linked immunosorbent assay (ELISA) and immunoblot assay.This demonstrates that recombinant acidic protein p9 has very good antigenicity and specificity and may be used for research on the in vitro and in vivo replication and immune responses in the vaccinated horses of donkey leukocyte-attenuated vaccine strain of equine infectious anemia virus.;