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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。
Human immunodeficiency virus (HIV) is one of the leading causes of AIDS. Gag, the core structural protein of HIV, has the ability to self-assemble and can stimulate humoral and cellular immunity, making it an ideal HIV vaccine target antigen. The Gag gene was cloned into the plasmid p Bac PAK8 to construct a recombinant plasmid, which was co-transfected into BmN cells with a linearized silkworm baculovirus genomic DNA to obtain the recombinant silkworm baculovirus Bm NPV-Gag. The recombinant virus infected Bm N cells and 5th instar larvae of silkworm, SDS-PAGE, Western blotting showed that Gm protein size (55 kD) was consistent with that of the recombinant virus infected Bm N cells and the 5th instar larvae of the silkworm The specific bands appeared to prove that the recombinant Gag protein was successfully expressed. The expression of target protein was detected by ELISA at different time points after infection with recombinant virus. The Gag protein content in Bm N cells reached the maximum of 6.995 pg / day on the fourth day after infection, and then decreased on the fifth day after infection. The Gag protein content in larval haemolymph was highest at 7 days after infection, at 790.1 ng / m L. The successful expression of recombinant Gag protein in the silkworm baculovirus expression system has explored new ways for the research of AIDS vaccine.