[Soluble expression and characterization of the immunoreactive multiepitope antigen of Toxoplasma gondii in E.coli].
OBJECTIVE
To obtain soluble expression product of immunoreactive recombinant multiepitope antigen of Toxoplasma gondii from E.coli.


METHODS
The gene encoding the multiple epitopes (MEG) of Toxoplasma gondii was amplified by PCR from the original plasmid containing MEG gene and cloned into the prokaryotic soluble expression vector pET32a. After identification by enzyme digestion and sequencing, the positive recombinant plasmid pET32a-MEG was transformed into BL21(DE3), which was induced with IPTG for expression of the target antigen. The relative molecular mass, solubility and antigenicity of the expression products were analyzed by SDS-PAGE and Western blotting.


RESULTS
The recombinant expression plasmid pET32a-MEG was successfully constructed and the highly efficient expression of the antigen was achieved after IPTG induction of E.coli. Improvement of the induction condition increased the expression product which accounted for about 28% of the total bacterial protein. The target protein, with good solubility and a relative molecular mass of about 31 000, was purified by immobilized metal affinity chromatography (Ni-NTA resin) and could be well recognized by mouse and rabbit antisera derived by infection of the animals with Toxoplasma gondii B36 and RH, respectively.


CONCLUSIONS
The recombinant multiepitope antigen has good antigenicity and potential value in diagnosis and vaccine development of toxoplasmosis.
Yang
Pei-liang
PL
Experimental Animal Center, Nanfang Hospital, Southern Medical University, Guangzhou 510515, China. plyang927@hotmail.com
Chen
Xiao-guang
XG
Wang
Yuan-zhan
YZ
Li
Hua
H
Zhou
Xiao-hong
XH
Wu
Kun
K
Yan
Hui
H
D000818:Animals; D000953:Antigens, Protozoan / Q000096:biosynthesis* / Q000235:genetics / Q000276:immunology; D003001:Cloning, Molecular; D000939:Epitopes; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D011817:Rabbits; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000235:genetics / Q000276:immunology; D014122:Toxoplasma / Q000276:immunology*; D014123:Toxoplasmosis / Q000276:immunology / Q000469:parasitology
D000953:Antigens, Protozoan; D000939:Epitopes; D011993:Recombinant Fusion Proteins