人类免疫缺陷病毒HIV结构蛋白Gag在家蚕杆状病毒表达系统中的表达

发布时间：2018-04-30 03:40

本文选题：人类免疫缺陷病毒 + Gag蛋白　；参考：《蚕业科学》2015年01期

【摘要】：人类免疫缺陷病毒(HIV)是导致艾滋病的主要病原之一,HIV病毒的核心结构蛋白Gag有自我装配功能,并能激发体液免疫和细胞免疫,是理想的HIV疫苗靶抗原。将Gag基因克隆至质粒p Bac PAK8中构建重组质粒,并与线性化的家蚕杆状病毒基因组DNA共转染Bm N细胞,得到重组家蚕杆状病毒Bm NPV-Gag。将该重组病毒感染Bm N细胞和家蚕5龄幼虫,SDS-PAGE、Western blotting检测显示在重组病毒感染的Bm N细胞和家蚕5龄幼虫血淋巴中均有与Gag蛋白大小(55 k D)相符的特异性条带出现,证明重组Gag蛋白成功表达。用ELISA法测定重组病毒感染后不同时间目的蛋白质的表达变化:Bm N细胞中的Gag蛋白含量在感染后的第4天达到最大值6.995 pg/个,在感染第5天后出现下降趋势;5龄幼虫血淋巴中的Gag蛋白含量在感染后的第7天最高,为790.1 ng/m L。重组Gag蛋白在家蚕杆状病毒表达系统中的成功表达,为艾滋病疫苗的研究探索了新的途径。
[Abstract]:Human immunodeficiency virus (HIV) is one of the main pathogens leading to AIDS. Gag, the core structural protein of HIV, has the function of self-assembly and can stimulate humoral and cellular immunity. It is an ideal target antigen of HIV vaccine. The Gag gene was cloned into the plasmid p Bac PAK8 to construct the recombinant plasmid and co-transfected with the linearized genomic DNA of Bombyx mori baculovirus into BmN cells to obtain the recombinant baculovirus BmNPV-Gag. The results of SDS-PAGEG blotting analysis of BMN cells infected with recombinant virus and silkworm 5th instar larvae showed that there were specific bands consistent with the size of Gag protein in hemolymph of Bm N cells infected by recombinant virus and silkworm larvae at 5th instar. It was proved that the recombinant Gag protein was successfully expressed. ELISA assay was used to determine the changes of protein expression at different times after infection of recombinant virus. The content of Gag protein in the cell line reached a maximum of 6.995 pg/ on the 4th day after infection. The Gag protein content in the hemolymph of the 5th instar larvae was the highest at the 7th day after infection, which was 790.1 ng/m L. The successful expression of recombinant Gag protein in Bombyx mori baculovirus expression system provides a new approach for the study of AIDS vaccine.
【作者单位】：浙江理工大学生命科学学院生物化学研究所浙江省家蚕生物反应器和生物医药重点实验室;
【基金】：大学生科技创新项目(No.2014R406041,2013R406018) 浙江省自然科学基金项目(No.Y3110354) 国家高技术发展计划“863”项目(No.2012ZX09102301-009,2011AA100-603) 浙江省高校生物学重中之重(一级)学科项目
【分类号】：R512.91

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