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首页> 外文期刊>Current Microbiology: An International Journal >Construction and expression of a eukaryotic expression vector containing a fusion gene of the hantaan virus s gene and hsp70 gene
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Construction and expression of a eukaryotic expression vector containing a fusion gene of the hantaan virus s gene and hsp70 gene

机译：包含汉坦病毒s基因和hsp70基因融合基因的真核表达载体的构建和表达

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摘要

Hantavirus (HV) infection leads to a kind of severe systematic syndrome, hemorrhagic fever with renal syndrome (HFRS). Heat shock proteins (HSPs) can be used as adjuvants assisting soluble antigens to produce specific targets which can be attacked by cytotoxic T lymphocytes. For further research on HFRS vaccine, this study aimed to express Hantaan virus nucleocapsid protein (HTNV NP)-HSP70 fusion protein in COS-7 cells. First, an HTNV S gene encoding NP was amplified by PCR with a mutated termination code and cloned into eukaryotic expression vector pCDNA3.1(+), into which the full-length hsp70 gene had already been inserted, to form the S-hsp70 fusion expression vector pCDNA3.1(+)/S-hsp70. Then this recombinant plasmid was transfected into COS-7 cells by liposome, and eukaryotic expression of NP-HSP70 fusion protein was detected by immunocytochemistry and western blot. The results show that the eukaryotic expression vector pCDNA3.1(+)/S-hsp70 was successfully constructed and the NP-HSP70 fusion protein was effectively expressed in COS-7 cells. This study demonstrates that the NP-HSP70 fusion protein was expressed effectively from the pCDNA3.1(+)/S-hsp70 vector in a eukaryotic system and thus provides a basis for using this plasmid as a new DNA vaccine against HV infection.
机译：汉坦病毒（HV）感染导致一种严重的系统综合征，即肾综合征出血热（HFRS）。热休克蛋白（HSP）可用作佐剂，协助可溶性抗原产生可被细胞毒性T淋巴细胞攻击的特异性靶标。为了进一步研究HFRS疫苗，该研究旨在在COS-7细胞中表达汉坦病毒核衣壳蛋白（HTNV NP）-HSP70融合蛋白。首先，使用突变的终止码通过PCR扩增编码NP的HTNV S基因，并将其克隆到已经插入了全长hsp70基因的真核表达载体pCDNA3.1（+）中，形成S-hsp70融合体表达载体pCDNA3.1（+）/ S-hsp70。然后通过脂质体将该重组质粒转染到COS-7细胞中，通过免疫细胞化学和Western blot检测NP-HSP70融合蛋白的真核表达。结果表明，成功构建了真核表达载体pCDNA3.1（+）/ S-hsp70，并在COS-7细胞中有效表达了NP-HSP70融合蛋白。这项研究表明，NP-HSP70融合蛋白可在真核系统中从pCDNA3.1（+）/ S-hsp70载体中有效表达，从而为将该质粒用作抗HV感染的新DNA疫苗提供了基础。

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