摘要
为获得J亚群禽白血病病毒(ALV-J)的gp85蛋白,将ALV-J的gp85基因克隆至pFastBac-HTA供体质粒,将其转入DH10BacTM大肠杆菌感受态细胞,使gp85基因整合到Bacmid穿梭载体中,构建重组穿梭载体Bacmid-gp85。通过脂质体介导,将重组穿梭载体Bacmid-gp85转染Sf9昆虫细胞,获得重组杆状病毒rBacgp85。Western-blot和间接免疫荧光试验(IFA)鉴定结果表明ALV-J gp85蛋白在Sf9昆虫细胞中得到正确表达,表达的重组gp85蛋白分子量约为38 ku。ALV-J gp85重组蛋白在Sf9细胞中的正确表达为其功能研究和应用提供了良好的基础。
In order to obtain the gp85 protein of J subgroup of avian leucosis virus （ALV-J）, gp85 gene was amplified and cloned into pFastBae-HTA to construct the recombinant donor plasmid pFastBae-HTA-gp85. Then, the pFastBac-HTA-gp85 was transformed into DH10Bac Escherichia coli competent cells to get the recombinant shuttle plasmid Bacmid-gp85. The recombinant baculovirus Bacmid-gp85 was obtained by transfecting rBac-gp85 with Cellfectina Reagen into St9 cells. The Western-blot analysis and indirect immunofluorescence assay revealed that the recombinant protein with the molecular weight of 38 kDa was expressed in the Sf9 cells. This study provides a good basis for functional analysis of gp85 protein of ALV-J.
出处
《畜牧与兽医》
北大核心
2011年第8期4-7,共4页
Animal Husbandry & Veterinary Medicine
基金
现代农业产业技术体系建设专项资金资助(nycytx-42-G3-01)
哈尔滨市科技攻关计划项目(2010AA6AN034)
