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| CCHFV S基因的重组杆状病毒的构建及鉴定 |
| Construction and identification of recombinant baculovirus with CCHFV S gene |
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| DOI：10.3969/j.issn.1007-8134.2020.05.005 |
| 中文关键词: 克里米亚-刚果出血热病毒 杆状病毒表达系统 核衣壳蛋白 |
| 英文关键词: Crimean-Congo hemorrhagic fever virus baculovirus expression vector systems nucleoprotein |
| 基金项目:陕西省社会发展科技攻关项目（2016SF-128）；空军军医大学军事医学提升计划（2016TSB-013） |
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| 中文摘要: |
| 目的 利用杆状病毒表达系统（baculovirus expression vector systems, BEVS）构建含有克里米亚-刚果出血热病毒（Crimean-Congo hemorrhagic fever virus, CCHFV）S基因的重组杆状病毒，并在sf9昆虫细胞中表达出核衣壳蛋白（nucleoprotein, NP）。方法 将合成的CCHFV S基因定向克隆入杆状病毒转移载体pFastBac? Dual，获得重组的转移载体pFastBac? Dual-CCHFV-S。将其转化到E. coli DH10Bac?感受态细胞中，筛选得到重组杆状病毒杆粒rBV-Bacmid-S。用Cellfectin? II试剂将rBV-Bacmid-S转染入sf9昆虫细胞获得重组杆状病毒rBV-CCHFV-S。通过IFA和Western blot检测CCHFV S基因的表达。结果 构建了含有S基因的rBV-CCHFV-S，在sf9昆虫细胞中成功表达CCHFV NP。结论 利用BEVS 可以成功表达CCHFV NP，这为研究CCHFV NP的生物学特性，研制特异性的治疗药物和预防控制疫苗奠定基础。 |
| 英文摘要: |
| Objective To construct recombinant baculovirus containing the S gene of Crimean-Congo hemorrhagic fever virus (CCHFV) is constructed using baculovirus expression vector systems (BEVS) and express nucleoprotein (NP) in sf9 insect cells. Methods The recombinant CCHFV S gene was directionally cloned into the baculovirus transfer vector pFastBacTM Dual to obtain the recombinant transfer vector pFastBacTM Dual-CCHFV-S, which was transformed into E. coli DH10BacTM competent cells and screened the recombinant bacmid rBV-Bacmid-S. Recombinant baculovirus rBV-CCHFV-S was obtained by transfecting rBV-Bacmid-S into sf9 insect cells with Cellfectin? II reagent. The expression of the CCHFV S gene was detected by IFA and Western blot. Results rBV-CCHFV-S containing S gene was constructed and CCHFV NP was successfully expressed in sf9 insect cells. Conclusions By using BEVS, CCHFV NP can be successfully expressed, which lays a foundation for the study of biological characteristics of CCHFV NP and develops of specific therapeutic drugs and preventive vaccines. |
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