Generation and characterization of a potentially applicable Vero cell line constitutively expressing the Schmallenberg virus nucleocapsid protein

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• 作者：Yongning Zhang ; Shaoqiang Wu ; Shanshan Song ; Jizhou Lv ; Chunyan Feng…
• 关键词：Schmallenberg virus ; Nucleocapsid protein ; Vero cells ; Lentivirus transduction ; Protein purification ; SBV bovine antisera
• 刊名：Cytotechnology
• 出版年：2017
• 出版时间：February 2017
• 年：2017
• 卷：69
• 期：1
• 页码：145-156
• 全文大小：
• 刊物类别：Chemistry and Materials Science
• 刊物主题：Biotechnology; Biomedicine, general; Biochemistry, general;
• 出版者：Springer Netherlands
• ISSN：1573-0778
• 卷排序：69

文摘

Schmallenberg virus (SBV) is a Culicoides-transmitted orthobunyavirus that poses a threat to susceptible livestock species such as cattle, sheep and goats. The nucleocapsid (N) protein of SBV is an ideal diagnostic antigen for the detection of viral infection. In this study, a stable Vero cell line, Vero-EGFP-SBV-N, constitutively expressing the SBV-N protein was established using a lentivirus system combined with puromycin selection. This cell line spontaneously emitted green fluorescent signals distributed throughout the cytoplasm, in which the expression of SBV-N fusion protein was confirmed by western blot analysis. The expression of SBV-N protein in Vero-EGFP-SBV-N cells was stable for more than fifty passages without puromycin pressure. The SBV-N fusion protein contained both an N-terminal enhanced green fluorescent protein (EGFP) tag and a C-terminal hexa-histidine (6 × His) tag, by which the N protein was successfully purified using Ni–NTA affinity chromatography. The cell line was further demonstrated to be reactive with SBV antisera and an anti-SBV monoclonal antibody in indirect immunofluorescence assays. Taken together, our results demonstrate that the Vero-EGFP-SBV-N cell line has potential for application in the serological diagnosis of SBV infection.