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Updated: Aug 9, 2026
Genetic Manipulation in Δku80 Strains for Functional Genomic Analysis of Toxoplasma gondii
Published on: July 12, 2013
[Expression and purification of Toxoplasma gondii GRA4 gene in prokaryotic system]
Qi-ping Lin1, Shao-ting Wu, Ya-biao Weng
1Shenzhen Center for Disease Control and Prevention, Shenzhen 518020, China.
Objective:
To construct a prokaryotic expression system containing the dense granule protein 4 (GRA4) of Toxoplasma gondii, purify the expressed protein and detect its immunogenicity.
Methods:
The specific fragment of GRA4 gene was amplified by PCR. After subcloning the prokaryotic expression recombinant pET-GRA4, the expressed product was purified with His Bind affinity chromatography and analyzed by Western blot. BALB/c mice were immunized with the GRA4 recombinant protein, and the antibody IgG titer was detected by ELISA.
Results:
The pET-GRA4 prokaryotic expression system was obtained. The MW of the expressed protein was Mr 40,000 and formed in inclusion body. After purification, the recombinant protein could be specifically recognized by the T. gondii infected rabbit serum. Mice immunized with the purified recombinant protein elicited high titer of IgG antibody.
Conclusion:
The pET-GRA4 recombinant protein was successfully expressed and purified, which shows the immunogenicity.
