新疆羊戊型肝炎血清流行病学调查及HEV ORF2-62基因的克隆表达

来源 :石河子大学 | 被引量 : 0次 | 上传用户:mywindjs
下载到本地 , 更方便阅读
声明 : 本文档内容版权归属内容提供方 , 如果您对本文有版权争议 , 可与客服联系进行内容授权或下架
论文部分内容阅读
HEV (Hepatitis E, HE), which can be a contagious zoonoses to animals and human beings caused by the hepatitis E virus. It is mainly spreaded from the feces to mouth, usually after the source of drinking water being contaminated; it will cause an outbreak of epidemic. There were sporadic cases globally. It had ever been occurred the world’s largest epidemic of hepatitis E in Hetian, Kashi and Kezilesu Kerkezi Autonomous State, which in the south of Xinjiang of China, in the eighties of last century. In addition, more and more data show that, HEV not only spread in humans, is also widely distributed in a variety of animals. In a wide range of animals from all over the world exist the natural HEV infection, and HEV isolated from animal and human had very high similarity, while human HEV have been successfully infected on a variety of animals experimentally. Islam as a faith-based religion in mainly minorities living in the Xinjiang region, the sheep is not only the most important livestock, but also the livestock closest contacted with the production and with people’s daily lives. So it not only has great significance to handle the epidemiological situation of sheep for human’s public health, but also to explore the host range of viruses, with little incidence of infected animals, how to spread between species and so many unknown issues. HEV antibodies were detected with double antigen sandwich enzyme-linked immunosorbent assay (DS-ELISA) for clarifing the HEV infection distribution on sheep in the different regions of Xinjiang and recombination cloning vector HEVORF2-62 and pET-28a prokaryotic expression vector had successfully been constructed, so it established the foundation for early indirect ELISA diagnostic method to HE.In order to learn prevalence situation of sheep hepatitis E virus infection in Xinjiang, we start this experiment on HEV.1131 sheep sera from 14 regions of Xingjiang (357 from Southern Xinjiang, 774 from Northern Xinjiang) were tested with DS-ELISA for the presence of anti-HEV IgG, and the prevalence differences were analysed based on location(South and North Xinjiang),age range(≤lyear,2-3years,≥3years) and species of sheep by X2 test. The result showed the overall prevalence of anti-HEV was 5.22%(59/1131), and 4.76%(17/357) for the South Xinjiang,and had no significant differences (P>0.05) than that of the North 5.42%(42/774). The prevalence rate of Sheep in spring (1.53%(7/458)) was significantly higher (P<0.05) than that of the Sheep in the fall (7.73%(52/673)). The result showed the sheep of Xinjiang has HEV in generally.In this paper, pig source ORF2-fusion gene, and express this in pET-28a prokaryotic expression vector were recombinated. Two pairs of degenerate primers were designed according to the genomic sequence of HEV landed submitted on Gene bank, ORF-62 gene 187bp was cloned by RT-PCR technology, and recombinatedwhich would be cloned into vector pBS-T, thus positive clones were got, then after sequencing, ORF-62 fusion gene was successfully constructed, named T-ORF-62.The vector T-ORF-62 and pET-28a were excised by BamHI, Hindlll respectively.225bp fragment and 5.3Kb vector fragment Recycling, then using T4 DNA ligase connected this two fragments to construct a recombinant prokaryotic expression plasmid pET-ORF-62. Through digestion and sequencing, the correct reading frame and the positive recombinant plasmid was obtaine.This recombinant plasmid was transformed into DE3 host expressing bacteria by IPTG,which has four density (0.8mM、1.0mM、1.2mM、1.4mM), for inducing and expressing then for sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Dot blot hybridization showed that expression of recombinant strains under the IPTG density of 1.2mM can successfully express ORF-62 fusion protein.
其他文献
当今社会发展迅速,人们的旅游需求不断增加.大量的车辆造成道路交通拥堵,在道路上造成一系列的交通问题,影响道路交通安全.城镇规模不断扩大,中小城镇快速发展,城市结构和规
文章主要分析了直缝埋弧焊管生产之中常见缺陷的形成原因和预防的对策.希望通过本文的分析,可以对直缝埋弧焊管生产缺陷几率的降低和生产质量的保障提供一定的参考.
路桥施工项目管理应用精细化管理,不但能够提升管理水平,有效控制施工成本,也能够保证路桥施工质量,使施工单位达成经济效益与社会效益的“双赢”.本文阐述了精细化管理的内
地基是公路的重要组成部分,在进行道路的施工过程中,软土地基的处理是一个非常重要的环节.软土地基施工技术的科学合理选择是保证公路施工质量和效率的关键,是提高公路使用性
RFID技术就是我们所说的无线射频识别技术.凭借着强大的自动识别能力,该新兴技术已经在我国的各个领域之中得到了广泛应用.本文就是对该技术在ETC(不停车收费)系统之中的应用
在公路的建设中离不开对软土地基技术的合理选取,应对复杂多样的软土地基类型时,针对性的拿出对应措施才能更精确的完成公路工程,从而使施工时间得到缩减的同时也为交通运输
期刊