Over-expression and purification of the recombinant p30 antigen of Toxoplasma gondii
10.3969/j.issn.1002-2694.2005.12.015
- VernacularTitle:刚地弓形虫P30(SAG1)重组抗原的高效表达和体外纯化
- Author:
Dianbo ZHANG
;
Defu ZAI
;
Maoqing GONG
;
Jin LI
;
Qingkuan WEI
;
Yong CUI
;
Bingcheng HUANG
;
Keyi LIU
- Publication Type:Journal Article
- Keywords:
T. gondii;
P30 gene;
fusion protein purification
- From:
Chinese Journal of Zoonoses
2005;(12):1089-1093
- CountryChina
- Language:Chinese
-
Abstract:
To provide the basis for preparation of diagnostic kits and vaccines in Toxoplasma gondii infection, the gene coding for the qualified recombinant p30 protein (SAG1) of this parasite was amplified by PCR, and the amplified gene was cloned into prokaryotic expression vector pET-30a(+) to construct the recombinant plasmid, and then transformed to E.coli DH5α. The positive recombinant plasmid was screened by PCR and double enzymes digestion, and the nucleotide sequence of p30 gene was determined by automated DNA sequencing. Meanwhile, the identified recombinant plasmid was transformed to E.coli BL21(DE3) with the expression of p30 on bacteria induced by IPTG and the expressed protein was identified by SDS-PAGE. The protein obtained was then further purified and refolded, and its biological activity was checked by Western blotting. It was shown that the size of the amplified gene was 750 bp with molecular weight of 30 ku, and this protein could specifically react with monoclonal antibody against p30 protein.
