Expression and Antigenicity Analysis of NTPase Gene of Toxoplasma gondii
- VernacularTitle:弓形虫三磷酸核苷水解酶基因克隆、表达与鉴定
- Author:
Dan SHA
;
Feng TAN
;
Changwang PAN
;
Shaohui LIANG
- Publication Type:Journal Article
- Keywords:
Toxoplasma gondii;
Nucleoside triphosphate hydrolase (NTPase);
Expression;
Antigenicity
- From:
Chinese Journal of Parasitology and Parasitic Diseases
1997;0(06):-
- CountryChina
- Language:Chinese
-
Abstract:
Objective To clone and express prokaryotic recombinant plasmid of nucleoside triphosphate hydrolase (NTPase) gene of Toxoplasma gondii, and analyze its antigenicity. Method NTPase gene was amplified by PCR from RH strain of T. gondii and cloned into pGEM-T Easy vector. Positive clones were screened and identified by BglⅡ, HindⅢ digestion and sequenced. The target gene was then subcloned into prokaryotic expression vector pBAD-HisB and transformed into E. coli BL21(DE3). The expressed recombinant protein was purified with Ni-NTA agarose and further analyzed by sodium dodecyl sulfate polyacrylamide gel electropheresis (SDS-PAGE) and Western blotting. Results NTPase-Ⅱ gene was specifically amplified, and the homology of DNA sequence was 100% to that in the GenBank. SDS-PAGE showed that the recombinant NTPase protein with correct molecular weight was expressed highly in E.coli BL21(DE3). Western blotting testified that the purified recombinant protein could be specifically recognized by mouse serum immunized with T. gondii and mouse anti-recombinant protein serum. Conclusion The NTPase-Ⅱ gene has been cloned and expressed in E.coli BL21(DE3), and the purified protein of NTPase-Ⅱ gene displays a specific antigenicity.
