Construction of single-chain fragment variable recombinant gene against SARS-CoV-2 S protein
10.16753/j.cnki.1008-2344.2025.01.006
- VernacularTitle:抗新型冠状病毒S蛋白scFv重组基因的构建
- Author:
Taoyuan YOU
1
;
Guangyong YANG
1
;
Guangzhi HE
1
Author Information
1. 贵州中医药大学基础医学院预防医学教研室,贵州 贵阳 550025
- Publication Type:Journal Article
- Keywords:
SARS-CoV-2;
spike protein;
single-chain fragment variable
- From:
Journal of Shenyang Medical College
2025;27(1):32-37
- CountryChina
- Language:Chinese
-
Abstract:
Objective:To construct a recombinant gene of single-chain fragment variable(scFv)against the SARS-CoV-2 spike protein(S protein).Methods:Using genetic engineering antibody technology,BALB/c mice were immunized with SARS-CoV-2 S protein antigen.The total RNA of the spleen was extracted to obtain the heavy chain(VH)and light chain(VL)gene sequences of the antibody variable region(V).Through multiple-primer PCR amplification and screening,the corresponding VH and VL gene sequences were obtained.After purification and recovery,the VH and VL primer genes of anti-SARS-CoV-2 S protein with restriction enzyme cutting sites and the VH and VL genes were successfully connected by the overlap extension PCR method to obtain the scFv recombinant gene.The sequencing results were compared and analyzed in the gene bank by using BLAST on the NCBI website.Results:The VH and VL genes of anti-SARS-CoV-2 S protein were successfully obtained,and scFv was obtained by connecting through the overlap extension PCR technique(SOE-PCR).The BLSAT analysis of scFv showed that its gene homology was greater than 80%.The indirect ELISA detection results showed that there was obvious binding in the prokaryotic expression supernatant.Conclusion:This study provides a technical basis for the diagnosis and treatment of SARS-CoV-2 and the research and development of new related antibodies.
