摘要
为获得鸭坦布苏病毒(duck Tembusu virus,DTMUV)E蛋白膜外区的表达产物,以DTMUV安徽分离株(AH10)E基因序列设计特异性引物,扩增E基因,将其克隆至原核表达载体pET-SUMO中,构建重组表达质粒pET-SUMO-E,然后将其转化至感受态细胞E.coli Rosetta(DE3)中,并以IPTG诱导表达。经SDS-PAGE检测,获得了70ku的表达产物,与预期结果相符。Western-blot分析表明,目的蛋白能与兔抗DTMUV E蛋白多克隆血清及DTMUV鸭阳性血清发生特异性反应,显示出良好的反应活性。结果表明,成功表达了E蛋白膜外区,这一成果为DTMUV疫苗制备和诊断试剂盒的研发奠定了基础。
According to the sequence of E gene of duck Tembusu virus AH10 strain,a primer pair was designed to amplify the ectodomain of E gene by PCR.The E gene was subsequently cloned into pET-SUMO vector.The resultant plasmid pET-SUMO-E was transformed into E.coli Rosetta（DE3） competent cells.The highly expressed product induced by IPTG was confirmed to be about 70 ku in size by SDS-PAGE,which was consistent with the expectation.Western-blot assay showed that it could react with both the rabbit anti-DTMUV polyclonal serum and the positive serum from DTMUV-infected duck. In summary,a high-efficient expression E gene in this study paves a way for development of new vaccines or diagnostic reagents.
出处
《中国兽医科学》
CAS
CSCD
北大核心
2013年第8期850-854,共5页
Chinese Veterinary Science
基金
甘肃省高层次人才科技创新创业扶持行动项目(1013JHTA008)
中央级公益性科研院所基本科研业务费专项(0032011027)
