Cloning and expression of gene encoding the mature peptide of microneme protein MIC3 from Toxoplasma gondii RH straimn

To clone the gene encoding the mature peptide of microneme preotein MIC3 from Toxoplasma gondii RH strain, constract its prokaryotic expression vector pET-MMIC3 to express on E.coli BL21 strain in order to investigate the mechanism of adhesion of T.gondii and its invasion to the host cells as well as the role of the MIC3 protein in these processes, the sequence encoding the mature microneme protein of MIC3 was amplified by PCR by using the genomic DNA from the tachyzoites of T.gondii RH strain as template.The amplified product was cloned into T-vector, identified and then subcloned into expression vector pET-30a(+), thus obtaining a recombinant plasmid pET-MMIC3. This recombinant vector was then transferred to E.coli BL21 strain (DE3). The induction was optimized, and a recombinant protein of 37.2 kDa was obtained, and the result of Western blotting was consistent with the calculated molecular weight. A prokaryotic recombinant plasmid to express the fusion protein of mature MIC3 was successfully constructed in the present study, and these results may provide the foundation for the further studies on the mechanism of T.gondii adhesion and invasion to host cells as well as the role of MIC3 in these processes.