摘要
为构建表达Ⅰ型鸭病毒性肝炎病毒(DHV-Ⅰ)VP1蛋白的重组腺病毒,本实验采用RT-PCR方法扩增DHV-Ⅰ HP-1株VP1基因,克隆于腺病毒穿梭质粒pShuttle-CMV中,并将其电转化于含腺病毒pAdeasy骨架质粒的大肠杆菌BJ5183中,通过同源重组制备含有重组腺病毒的质粒。重组质粒经PacⅠ酶切线性化后转染AD293细胞,获得重组腺病毒(rAd-VP1)。间接免疫荧光和western blot鉴定结果显示,DHV-Ⅰ的VP1蛋白在重组腺病毒感染的AD293中获得表达。rAd-VP1的制备为鸭免疫实验效果评价奠定了基础。
In order to construct recombinant adenovirus expressing the VP1 protein of duck hepatitis virus type I （DHV-I）, the VPI gene was amplified by RT-PCR and cloned into pShuttle-CMV vector. The recombinant plasmid was transferred into E. co/i BJ5183 competent cells containing adenovirus backbone vector to produce recombinant adenovirus plasmid by homologous recombination. The positive recombinant plasmid was linearized by Pac I and transfected into AD293 cells. The recombinant adenovirus （rAd-VPI） was rescued and identified by indirect immunofluorescence assay and westem blot. The results showed that rAd-VPl expressing VPI protein of DHV- I was constructed successfully which laid the foundation for the evaluation of immune efficacy in ducks.
出处
《中国预防兽医学报》
CAS
CSCD
北大核心
2013年第7期526-529,共4页
Chinese Journal of Preventive Veterinary Medicine
基金
现代农业水禽产业技术体系岗位科学家专项(CARS-43-10)
