Related Experiment Video
Updated: May 7, 2026
Small-Scale Plasma Membrane Preparation for the Analysis of Candida albicans Cdr1-mGFPHis
Published on: June 13, 2021
[Prokaryotic expression and immunogenicity analysis of phosphoglycerate kinase 1 of Candida albicans]
Zhengxin He1, Jing Chen, Xiangyang Ran
1Department of Clinical Laboratory, Bethune International Peace Hospital of PLA, Shijiazhuang 050082, China.
Insights
Researchers successfully expressed the Candida albicans phosphoglycerate kinase 1 (PGK-1) protein using a prokaryotic system. The resulting recombinant PGK-1 protein demonstrated significant immunogenicity in mouse models.
Area of Science:
- Molecular Biology
- Immunology
- Microbial Genetics
Background:
- Candida albicans is a significant human fungal pathogen.
- Phosphoglycerate kinase 1 (PGK-1) is a crucial enzyme in glycolysis.
- Understanding C. albicans PGK-1 can aid in developing diagnostic and therapeutic strategies.
Purpose of the Study:
- To construct prokaryotic expression plasmids for Candida albicans phosphoglycerate kinase 1 (pgk-1).
- To express and purify the recombinant PGK-1 protein.
- To evaluate the immunogenicity and antigenicity of the recombinant PGK-1 protein in a murine model.
Main Methods:
- Amplification and cloning of the full-length pgk-1 coding sequence into the pET30a vector.
- Induction of 6×His-tagged recombinant protein expression in E. coli BL-21(DE3).
- Identification of the recombinant protein via SDS-PAGE and Western blotting.
- Immunization of BALB/c mice with purified recombinant PGK-1 and antibody titer determination by ELISA.
Main Results:
- Successful construction of the pET-30a-pgk-1 expression plasmid.
- High-level expression of the recombinant PGK-1 protein (approx. 54.8 kDa) in E. coli.
- ELISA confirmed favorable immunogenicity, with antibody titers reaching approximately 1:1024.
Conclusions:
- The prokaryotic expression system efficiently produced recombinant Candida albicans PGK-1.
- The recombinant PGK-1 protein elicits a robust immune response in mice.
- This study provides a foundation for further investigation into PGK-1 as a potential diagnostic or vaccine target.
Objective:
To construct prokaryotic expression plasmids of Candida albicans gene phosphoglycerate kinase 1 (pgk-1) and examine the immunogenicity of the recombinant protein.
Methods:
The full-length coding sequence of pgk-1 was amplified by PCR and cloned into the prokaryotic expression vector pET30a. The 6×His-tagged protein was induced by IPTG in E.coli BL-21(DE3) and the recombinant protein was identified by SDS-PAGE and Western blotting. The BALB/c mice were immunized with the purified recombinant protein to evaluate the antigenicity of the recombinant protein by ELISA.
Results:
The full-length pgk-1 gene was cloned from SC5314 genome and pET-30a-pgk-1 was successfully constructed. The recombinant protein PGK-1 was highly expressed in E.coli with a relative molecule mass of 54 810. ELISA indicated that the titer of the antibody was about 1:1024.
Conclusion:
PGK-1 was successfully expressed by prokaryotic expression system and the recombinant protein showed favorable immunogenicity in mice.
