[High efficiency expression and antigenicity analysis of the SAG2 gene from Toxoplasma gondii RH strain].
OBJECTIVE
To make high efficiency expression of the SAG2 gene from Toxoplasma gondii RH strain in E. coli and study the antigenicity of the expressed product.


METHODS
The SAG2 gene fragment of T. gondii RH strain amplified by PCR method from genome DNA was cloned into the pMD-18T vector and transformed into E. coli DH5alpha. After nucleotide sequencing, the SAG2 gene fragment was subcloned into the expression vector pET23a with correct orientation and transformed into E. coli DH5alpha. The plasmid from the correct clone identified by PCR method and endonuclease digestion was transformed into E. coli BL21 (DE3) and induced for expression. The expressed product was studied by SDS-PAGE and Western blot.


RESULTS
502 bp gene fragment was amplified by PCR as anticipated. Nucleotide sequencing showed a 100% homology with that of the published sequence in GenBank. The molecular weight of the expressed protein was about Mr 19,000. Western blotting indicated that the antigenicity of the protein was specific.


CONCLUSIONS
The plasmid pET23a-SAG2 was constructed and a high efficiency expression of the SAG2 gene from T. gondii RH strain was made. The expressed product shows a specific antigenicity.
Lei
Ming-jun
MJ
Tongji Medical College of Huazhong University, Wuhan 430030, China.
Wu
Shao-ting
ST
Dai
Wu-xing
WX
Pan
Hui-rong
HR
Lin
Qi-ping
QP
Wen
Jian-xiang
JX
Huang
Da-na
DN
Gao
Shi-tong
ST
Zhang
Ren-li
RL
D000818:Animals; D000953:Antigens, Protozoan / Q000096:biosynthesis* / Q000235:genetics / Q000276:immunology*; D015153:Blotting, Western; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D006801:Humans; D015800:Protozoan Proteins / Q000096:biosynthesis* / Q000235:genetics / Q000276:immunology*; D014122:Toxoplasma / Q000145:classification / Q000235:genetics / Q000276:immunology*
D000953:Antigens, Protozoan; D015800:Protozoan Proteins; C067547:surface antigen P22, Toxoplasma