在昆虫细胞中使用快速新的表达系统生产具有免疫反应性的多瘤病毒和疱疹病毒重组糖蛋白,用于疫苗和血清学检测

B. Abedi Kiasari, H. Najafi
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引用次数: 0

摘要

引言:尽管用于杆状病毒血清学检测和疫苗开发的多瘤病毒和疱疹病毒重组蛋白的传统表达方法已经建立,但操作既费力又耗时。方法:利用一种新的质粒表达系统在昆虫细胞中表达两种多瘤病毒主要衣壳蛋白VP1(JCV VP1和BKV VP1)和两种疱疹病毒糖蛋白(HSV-1 gD和VZV gE)。采用不依赖连接的克隆法(LIC)构建重组质粒。使用重组体进行Sf9昆虫细胞的转染。使用SDS-PAGE、免疫荧光和免疫印迹分析产生的蛋白质。结果:JCV-VP1、BKV-VP1,VZV-gE和HSV-1gD在感染后48h在昆虫细胞中成功表达,并在细胞质和细胞膜中检测到免疫反应性。这种基于质粒的表达系统需要5天的时间来表达蛋白质。结论:基于质粒的昆虫细胞表达系统是高效的,是在昆虫细胞中快速表达多瘤病毒和疱疹病毒蛋白的理想系统,有可能用于疫苗成分和血清学检测等应用。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
Production of Polyomavirus and Herpesvirus Recombinant Glycoproteins with Immunoreactivity Using a Rapid and Novel Expression System in Insect Cells for Applications in Vaccines and Serological Assays
Introduction: Although, conventional methods for the expression of polyomaviruses and herpesviruses recombinant proteins for serological assays and vaccine developments in baculoviruses are well established, the manipulations are laborious and time consuming. Methods: A new expression system based on plasmid was used to express two polyomaviruses major capsid protein VP1 (JCV VP1 and BKV VP1), and two herpesviruses glycoproteins (HSV-1 gD and VZV gE) in insect cells. A ligation independent cloning (LIC) was applied to generate the recombinant plasmids. Transfection of Sf9 insect cells were performed using the recombinants. The produced proteins were analysed using SDS-PAGE, immunofluorescence, and immunoblotting. Results: JCV-VP1, BKV-VP1, VZV-gE and HSV-1gD were successfully expressed in the insect cells, 48 h post-infection and detected in cytoplasm and cell membranes with immunoreactivity. This plasmid based expression system took 5 days to express the protein. Conclusion: The plasmid based expression system in insect cells was highly efficient and would be ideal for rapid expression of polyomaviruses and herpesviruses proteins in insect cells to be potentially used in applications such as vaccine components and serological assays.
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