Objective: To clone and express the phosphoglycerate mutase 2 (PGAM2) gene of Toxoplasma gondii, and analyze the antigenicity of the recombinant protein.
Methods: Total RNA was extracted from T. gondii tachyzoites of RH strain and reversely transcribed into cDNA. TgPGAM2 gene was amplified by PCR and cloned into pET30a(+) vector. The constructed pET30a(+)-TgPGAM2 was transformed into E. coli DH5alpha first and selected through the colony-PCR and confirmed by the double restriction enzyme digestion and sequencing. The correct plasmid was transformed into E. coli BL21 for expression induced by IPTG and the recombinant protein was further analyzed through SDS-PAGE followed by Coomassie brilliant blue staining. Western blotting assay with rabbit anti-T. gondii serum was used to analyze its antigenicity.
Results: The length of PCR product was about 750 bp and the recombinant plasmid pET30a(+)-TgPGAM2 was successfully constructed. The results of SDS-PAGE and Western blotting revealed that the relative molecular weight (Mr) of the soluble recombinant protein was approximately 30 000 and could be recognized by rabbit anti-T. gondii serum.
Conclusion: The soluble TgPGAM2 protein has been expressed in the prokaryotic expression system and maintains its antigenicity.
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