猪圆环病毒2型ORF1和ORF3 T淋巴细胞抗原表位的克隆与原核表达

摘要

根据Stevenson筛选出来的3个具有活性的T淋巴细胞抗原表位(T lymphocyte epitope,TCE)合成tce基因(180 bp),成功插入到pMD18-T Simple载体中,筛选获得重组质粒,命名为pMD-TCE.将tce酶切产物亚克隆到原核表达载体pET-32a(+)中,获得重组质粒,命名为pET-TCE.用IPTG进行诱导表达,收集菌液进行SDS-PAGE检测,结果显示合成基因在pET-32a(+)中获得了高效融合表达,其表达蛋白相对分子质量约为25 000,Western-blot分析结果表明,获得的融合蛋白可与猪圆环病毒2型(PCV2)阳性血清发生特异性反应,具有良好的免疫原性.%According to three active T lymphocyte epitopes ( TCE) which Stevenson screened, tee gene (180 bp) was synthesized and cloned successfully into pMD18-T Simple vector, tee gene was subcloned into prokaryotic expression vector pET-32a( + ); the recombinant plasmid was named pET-TCE and induced by IPTG. The results of SDS-PAGE and Western-blot indicated lhat the tce gene was expressed at a high level, and the recombinant fusion protein was about 25 000. The recombinant fusion protein has a peculiar reaction to porcine circovirus 2(PCV2) positive serum which has immunological activity.