Prokaryotic expression of human group A rotavirus VP6 protein and preparation of mouse polyclonal antibodies
10.3969/j.issn.1002-2694.2025.00.143
- VernacularTitle:人A组轮状病毒VP6蛋白的原核表达及其小鼠多克隆抗体的制备
- Author:
Weibin ZHANG
1
;
Changkun LYU
;
Ying YANG
;
Xiangpeng WANG
;
Mingyong WANG
Author Information
1. 商丘医学高等专科学校医学技术学院,商丘 476100
- Publication Type:Journal Article
- Keywords:
human group A rotavirus;
VP6 protein;
prokaryotic expression;
polyclonal antibodies
- From:
Chinese Journal of Zoonoses
2025;41(9):910-916
- CountryChina
- Language:Chinese
-
Abstract:
This study expressed the VP6 protein of human group A rotavirus in a prokaryotic expression system and prepared mouse polyclonal antibodies to this protein.The human rotavirus VP6 gene was amplified through RT-PCR from fecal samples infected with group A rotavirus and cloned into the pET-32a vector to construct the recombinant pET-32a-VP6 prokaryotic expression plas-mid.The recombinant plasmid was transformed into Escherichia coli BL21(DE3).VP6 protein was expressed through induction with isopropyl-β-D-thiopyranoside(IPTG),purified with nickel nitrilotriacetic acid(Ni-NTA)affinity chromatography,and identified through SDS-PAGE and western blotting.Polyclonal antibodies to the VP6 protein were obtained by immunization of BALB/c mice with VP6 protein.Antibody titers were determined through indirect ELISA.The recombinant expression plasmid pET-32a-VP6 was con-structed,and VP6 protein was expressed primarily in inclusion bodies.Polyclonal antibodies to VP6 protein with a titer of 1∶32 000 were obtained by immunization of mice with VP6 protein purified by affinity chromatography.The antibodies specifically reacted with VP6 protein.The VP6 protein of human group A rotavirus was expressed and purified,and mouse polyclonal antibodies to this protein were prepared,thus providing a foundation for the preparation of a diagnostic antigen for human group A rotavirus and the establishment of serological detection methods.
