[Cloning, expression and purification of the major surface antigen P30 of Toxoplasma gondii].
OBJECTIVE
To obtain protein of the major surface antigen P30 of Toxoplasma gondii by molecular cloning.


METHODS
The gene of P30 (containing the whole P30 gene sequence, without the gene encoding signal peptide) was obtained by polymerase chain reaction (PCR) using the primer designed according to the DNA sequence of P30. The recombinant plasmid was constructed using EcoR I, Xho I and was then transformed into E. coli Top10. The positive clones were identified by restriction enzymes and DNA sequence analysis. The fusion protein was induced by IPTG and purified by affinity chromatography using ProBond Resin (a kind of nickel-charged sepharose resin) and was identified by SDS-PAGE and Western blotting.


RESULTS
The products of PCR, cleavage and link reaction were same as expected and the sequence of inserted fragment in the recombinant plasmid was same as reported except one synonymy mutation. A 58 kDa fusion protein was induced by IPTG and was purified by chromatography.


CONCLUSIONS
Fusion protein containing Toxoplasma gondii P30 was achieved and was provided as experiment material for further research.
Zheng
Da-li
DL
Department of Biochemistry and Molecular Biology, Fujian Medical University, Fuzhou 350004.
Huang
Qing-ling
QL
Zhang
Tao
T
Lin
Jian-yin
JY
D000818:Animals; D000953:Antigens, Protozoan / Q000096:biosynthesis* / Q000235:genetics / Q000302:isolation & purification; D000954:Antigens, Surface / Q000096:biosynthesis* / Q000235:genetics / Q000302:isolation & purification; D015153:Blotting, Western; D003001:Cloning, Molecular; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000235:genetics; D016133:Polymerase Chain Reaction; D015800:Protozoan Proteins / Q000096:biosynthesis* / Q000235:genetics / Q000302:isolation & purification; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis* / Q000302:isolation & purification; D014122:Toxoplasma / Q000276:immunology*
D000953:Antigens, Protozoan; D000954:Antigens, Surface; D015800:Protozoan Proteins; D011993:Recombinant Fusion Proteins; C047168:SAG1 antigen, Toxoplasma