人博卡病毒样颗粒的研制及初步应用

发布时间：2018-04-02 17:23

本文选题：人博卡病毒　切入点：杆状病毒表达载体系统　出处：《武汉生物制品研究所》2010年硕士论文

【摘要】： 2005年，瑞典学者首次报道在呼吸道疾病患者鼻咽分泌物中发现人博卡病毒1，随后，人博卡病毒2和人博卡病毒3又分别在急性迟缓型麻痹患儿和腹泻患儿的粪便中被相继发现。人博卡病毒是一类无包膜的单链线状DNA病毒，暂时划归为细小病毒科、细小病毒亚科、博卡病毒属。由于缺乏有效的体外细胞培养体系和动物模型，人博卡病毒与人类呼吸道或胃肠道疾病的临床关联尚且未知。因此，使用基因工程方法大量制备人博卡病毒抗原，建立相应的血清学检测方法，将有助于解开这些疑问。本研究使用昆虫细胞和杆状病毒表达载体系统表达人博卡病毒1、2和3衣壳蛋白VP2基因，重组衣壳蛋白VP2部可以自行组装成病毒样颗粒，使用纯化的人博卡病毒样颗粒建立酶联免疫吸附试验，检测200份西安地区健康婴儿、176份北京地区健康人血清中人博卡病毒1、2和3的IgG和IgM水平，以及373份日本福冈地区健康人血清中人博卡病毒1的IgG水平。电泳及透射电镜结果显示，大量分子量约为60．5kD的蛋白释放入培养上清中，，重组蛋白可以自行组装成不含核酸的空心病毒样颗粒，这些病毒样颗粒平均直径为22nm，颗粒密度约为1．31g／ml。人博卡病毒血清流行病学结果显示，婴儿中人博卡病毒1、2和3的IgG阳性率分别为20．0％、14．0％和13．5％；而10岁以上人群中人博卡病毒1、2和3的IgG阳性率升高至75．6％、44．3％和42．0％；但是日本福冈地区人博卡病毒1的IgG阳性率超过80％。与IgG流行趋势相反，婴儿中人博卡病毒的IgM阳性率明显高于成年人群。昆虫细胞和杆状病毒表达载体系统可以有效表达人博卡病毒1、2和3衣壳蛋白VP2，并自行组装成病毒样颗粒。人博卡病毒在普通人群中广泛流行，成年人群中人博卡病毒的IgG的阳性率很高。
[Abstract]:In 2005, Swedish scholars first reported the discovery of human Boca virus 1 in nasopharyngeal secretions of patients with respiratory diseases. Subsequently, human Boca virus 2 and human Boca virus 3 were found in feces of children with acute delayed paralysis and diarrhea, respectively.Human Boca virus is a class of uncoated single strand linear DNA virus, which is classified as parvovirus family, parvovirus subfamily and Boca virus genus.Due to the lack of effective cell culture system in vitro and animal models, the clinical association between human Boca virus and human respiratory or gastrointestinal diseases is unknown.Therefore, it is helpful to solve these questions by using genetic engineering method to prepare human Boca virus antigen in large quantities and to establish corresponding serological detection method.In this study, the insect cell and baculovirus expression vector system was used to express the VP2 gene of human Boca virus 1 / 2 and 3 capsid protein. The recombinant capsid protein VP2 could be self-assembled into virus-like particles.Enzyme linked immunosorbent assay (Elisa) was established using purified human Boca virus-like particles to detect the levels of IgG and IgM in sera of 200 healthy infants from Xi'an area and 176 healthy people from Beijing area.The IgG level of human Boca virus 1 in 373 healthy people in Fukuoka, Japan.The results of electrophoretic and transmission electron microscopy showed that a large number of proteins with molecular weight of about 60.5kD were released into the supernatant, and the recombinant protein could be self-assembled into hollow virus-like particles without nucleic acid. The average diameter of these virus-like particles was 22 nm and the particle density was about 1.31 g / ml.The results of seroepidemiology of human Boca virus show thatThe positive rates of IgG in infants were 20.0% and 13.50.The IgG positive rates of human Boca virus 1and 3 increased to 75.643% and 42.0%, respectively, but the IgG positive rate of Boca virus 1 was more than 80% in Fukuoka area, Japan.Contrary to the epidemic trend of IgG, the positive rate of IgM in infants was significantly higher than that in adults.Insect cells and baculovirus expression vector system can effectively express human Boca virus 1 / 2 and 3 capsid protein VP2and self-assemble virus-like particles.Human Boca virus is prevalent in the general population, and the positive rate of human Boca virus IgG is very high in adult population.
【学位授予单位】：武汉生物制品研究所
【学位级别】：硕士
【学位授予年份】：2010
【分类号】：R373

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