Cloning, Expression and Identification of Surface Antigen SAG4 of Toxoplasma gondii

Objective To clone and express surface antigen SAG4 gene of Toxoplasma gondii, and analyze its immunoreactivity. Methods Specific primers were designed based on the reported SAG4 gene of T. gondii RH strain (GenBank Accession No: AF340224.1). Using genomic DNA from T. gondii as templates, SAG4 gene was amplified by PCR. The PCR product was cloned into pMD19-T vector and identified by digestion with restriction enzyme and PCR. Then the target fragment was subcloned into pET28a(+) vector, transformed into E. coli BL21 and followed by expression of the protein induced by IPTG. The protein was identified by Western blotting. Results The target gene was amplified with the length of 537 bp. Sequence analysis showed that the predicted amino acid sequence was identical with that of SAG4 as a membrane protein in T. gondii. After induced by IPTG, the recombinant SAG4 protein existed in an inclusion body form. The recombinant SAG4 (Mr 18 740) was recognized by serum of infected mice. Conclusion SAG4 has been expressed and shows certain immuno-response activity.