Prokaryotic expression of GII.4 human norovirus VP2 protein and preparation of anti-VP2 polyclonal antibody
10.3760/cma.j.cn112866-20220403-00083
- VernacularTitle:GII.4型人诺如病毒VP2蛋白的原核表达与多克隆抗体制备
- Author:
Yalin MA
1
;
Jindong WANG
;
Tongyao MAO
;
Qing ZHANG
;
Xiangyu KONG
;
Zhaojun DUAN
Author Information
1. 甘肃中医药大学公共卫生学院，兰州 730000
- Keywords:
Human norovirus;
VP2 protein;
Prokaryotic expression;
Polyclonal antibody
- From:
Chinese Journal of Experimental and Clinical Virology
2023;37(1):78-82
- CountryChina
- Language:Chinese
-
Abstract:
Objective:To express prokaryotically GII.4 human norovirus (HuNoV) VP2 protein and to prepare polyclonal antibody against VP2.Methods:Design specific primers to amplify the VP2 gene of GII.4 HuNoV, digest and connect to the prokaryotic expression vector pGEX-6P-1, transform the correctly identified recombinant plasmid into BL21 ( DE3) competent cells.Pick out and shake the monoclonal bacteria, and add IPTG to induce recombinant GST-VP2. The fusion protein was expressed, purified by GST affinity chromatography and digested to obtain GII.4 HuNoV VP2 protein. The relative molecular mass (Mr.×10 3) of the purified HuNoV VP2 protein was analyzed by SDS-PAGE. BALB/c mice were immunized with purified GII.4 HuNoV VP2 protein (0.5 mg/ml) to prepare polyclonal antibodies. Results:The VP2 protein of GII.4 HuNoV was successfully expressed and purified, with a relative molecular mass (Mr.×10 3) of about 29; the VP2 polyclonal antibody of GII.4 HuNoV was successfully prepared and its titer was as high as 1∶1 280 000. Western blot and indirect ELISA analysis showed that the polyclonal antibody could specifically bind to the VP2 antigen of GII.4 HuNoV. Conclusions:The purified GII.4 HuNoV VP2 after prokaryotic fusion expression can be used to prepare high titer polyclonal antibody.
