Abstract:

Objective   To construct the recombinant eukaryotic plasmid encoding rhoptry protein 18(ROP18) and microneme 8 (MIC8) of Toxoplasma gondii and identify it in the transcription and translation level. Methods   The recombinant eukaryotic expression plasmid pBudCE4.1-ROP18-MIC2 was constructed by the molecular cloning technology. After being identified by PCR, restriction enzyme cleavage and sequencing, pBudCE4.1-ROP18-MIC2 was transfected into the HFF cells.We then detected the transcription level at 48h using RT-PCR and examined the protein expression at 72h by SDS-PAGE. Results   Analysis of PCR, restriction enzyme cleavage and sequencing confirmed that the recombinant eukaryotic expression plasmid of pBudCE4.1-ROP18-MIC2  was precisely constructed.The identification of RT-PCR and the appraisal of SDS-PAGE showed that  ROP18 and MIC2  of  Toxoplasma gondii got the correct transient expression in the HFF cells. Conclusion   The successful constrution of the recombinant eukaryotic expression plasmid pBudCE4.1-ROP18-MIC2 has laid foundation for the further development of protective vaccine against Toxoplasma gondii infections.

Key words: Toxoplasma； Genes, ROP18; Genes, MIC2； Cloning, molecular； Eukaryotic cells