Clonging Expression And Identification The Rhoptry Protein2 (ROP2) Gene Fragment Of QHO Strains Of Toxoplasma Gondii

Toxoplasma gondii is an obligate intracellular parasite that infects a wide range of hosts, including humans and domesticated animals throughout the world, and it has been estimated that one-third of the human population is infected. Infection of Immunocompetent humans is usually asymptomatic, with clinical disease largely confined to risk groups. Primary infection during pregnancy can result in severe neonatal malformations and ocular complications in the fetus. Since the emergence of AIDS, toxoplasmosis in the immunocompromised host is of great significance, as the recrudescence of a latent infection often results in a fatal encephalitis. In addition, toxoplasmosis can cause considerable economic loss in the farming industry. Thus, the development of an effective vaccine against Toxoplasma gondii would be of great value to both human and veterinary medicine., due to ROP2 antigen is secreted by the rhoptry of tachyzoites and bradyzoites. They are highly conservative and immunogenic, were selected as antigens for diagnosis and vaccination totoxoplasmosis. To investigate cloning and expression of the ROP2 gene fragment of QHO strains of Toxoplasma gondii. ROP2 gene fragment was amplified by PCR from the genomic DNA of QHO strains of Toxoplasma gondii. the gene of ROP2 was inserted into the plasmid pMD18-T,and they was transferred into E.coli JM109. After the identification by PCR and restrictive digestion. Then the recombinant was sequenced, and the correct sequence of open reading fragment (ORF) was gained. Homology analysis showed that the ROP2 sequences of QHO strain shared 97.4% homology with that of RH strain, and the deduced amino acid sequences from ROP2 sequences of QHO strain shared 93.9%homology with that from ROP2 sequences of RH strain. The sequence of ROP2 compared with other rhoptry proteins ,the results showed that they perhaps have a commom origin which come from several protein familys.1603bp(Ql) and 1132bp(Q2) fragments of the ROP2 gene was amplified by PCR from the recombinant plasmid pMD18-T-ROP2 and The amplified fragments were subcloned into the polylinker of the plasmid pET-28b .Then the recombinant plasmids pET-Qlå’ŒpET-Q2 were transformated into E. coli BL21(DE3) The positive clones BL21(DE3) containing recombinants were idenditied by the methods of restrictive digestion and the gene sequencing. The recombinant E.coli were induced by IPTG to express the fusion protein. The expressed and purified products were analyzed by SDS-PAGE and Western-blot, this result will pave the way for the development of Toxoplasma gondii vaccine.