Protein expression, purification and mouse antiserum preparation of monkeypox virus A23R.
- Author:
Yihao WANG
1
;
Mingzhi LI
1
;
Mengle JIA
1
;
Lingdi YANG
1
;
Jiaqi XIONG
1
;
Ting WANG
1
;
Yu WANG
1
;
Shurong LIU
1
;
Wenli GUO
1
;
Lingbao KONG
2
;
Meifeng LI
3
Author Information
1. Institute of Pathogenic Microorganism, Nanchang City Key Laboratory of Animal Virus and Genetic Engineering, Department of biotechnology, College of Bioscience and Engineering, Jiangxi Agricultural University, Nanchang 330045, China.
2. Institute of Pathogenic Microorganism, Nanchang City Key Laboratory of Animal Virus and Genetic Engineering, Department of biotechnology, College of Bioscience and Engineering, Jiangxi Agricultural University, Nanchang 330045, China. *Corresponding authors, E-mail: lingbaok@mail.jxau.edu.cn.
3. Institute of Pathogenic Microorganism, Nanchang City Key Laboratory of Animal Virus and Genetic Engineering, Department of biotechnology, College of Bioscience and Engineering, Jiangxi Agricultural University, Nanchang 330045, China. *Corresponding authors, E-mail: meifengli77@jxau.edu.cn.
- Publication Type:Journal Article
- MeSH:
Animals;
Mice;
Monkeypox virus;
Antibodies;
Enzyme-Linked Immunosorbent Assay;
Blotting, Western;
Recombinant Proteins;
Escherichia coli/genetics*
- From:
Chinese Journal of Cellular and Molecular Immunology
2023;39(7):642-648
- CountryChina
- Language:Chinese
-
Abstract:
Objective To express the monkeypox virus (MPXV) A23R protein in Escherichia coli and purify by Ni-NTA affinity column, and to prepare mouse antiserum against MPXV A23R. Methods The recombinant plasmid pET-28a-MPXV-A23R was constructed and transformed into Escherichia coli BL21 to induce the expression of A23R protein. After optimizing the conditions of expression, A23R protein was highly expressed. Recombinant A23R protein was purified by Ni-NTA affinity column and identified by Western blot analysis. The purified protein was used to immunize mice for preparing the A23R polyclonal antibody, and the antibody titer was detected by ELISA. Results The expression of A23R recombinant protein reached the peak under the induced conditions of 0.6 mmol/L isopropyl-β-D-thiogalactoside (IPTG), 37 DegreesCelsius and 20 hours. The purity of the protein was about 96.07% and was identified by Western blot analysis. The mice were immunized with recombinant protein, and the titer of antibody reached 1:102 400 at the 6th week after immunization. Conclusion MPXV A23R is expressed highly and purified with a high purity and its antiserum from mouse is obtained with a high titre.
