人3型腺病毒六邻体基底区重组蛋白的克隆表达和纯化

Cloning,expression and purification of basal region fragment of hexon protein in human adenovirus

目的克隆、表达人腺病毒六邻体蛋白基底区片段,为后续研究六邻体蛋白的抗原性及蛋白结构,进而研发腺病毒的基因工程疫苗及诊断试剂奠定基础。方法以3型人腺病毒核酸为模板,用PCR法扩增六邻体基底区片段,将该片段定向克隆至p QE32质粒,在大肠杆菌表达系统中表达带有组氨酸标签的重组蛋白片段,利用镍亲和层析法对重组蛋白进行纯化。结果构建出重组表达质粒,通过测序验证了克隆结果的正确性;在大肠杆菌表达体系中获得了高效表达。结论成功表达并获得了纯化的人3型腺病毒六邻体基底区重组蛋白片段。

Objective To clone and express basal region fragment of hexon protein from HAdV-3 as to lay a foundation for thorough understanding the antigenicity and protein structure of hexon, which may lead to development of a vaccine candidate or an Adv diagnostic reagent. Methods The hexon protein basal region fragment was amplified by PCR using genomic DNA of HAdV-3 （human adenovirus 3 ） as template. Then the target DNA was directionally cloned to pQE32 plasmid. The recombinant plasmid was transformed into competent E. coli. M15 to express the target protein. Then the target protein was purified by Ni-Resin column affinity chromatography. Results The recombinant plasmid was successfully constructed. The recom- binant protein was highly expressed in E. coli. M15. Conclusion The fragment of HAdV-3 hexon protein basal region is successfully cloned and expressed.