Expression and activity determination of recombinant capsid protein VP2 gene of enterovirus type 71 / 中华预防医学杂志
Chinese Journal of Preventive Medicine
;
(12): 324-327, 2014.
Article
Dans Chinois
| WPRIM
| ID: wpr-298927
ABSTRACT
<p><b>OBJECTIVE</b>To clone and express the recombinant capsid protein VP2 of enterovirus type 71 (EV71) and to identify the immune activity of expressed protein in order to build a basis for the investigation work of vaccine and diagnostic antigen.</p><p><b>METHODS</b>VP2 gene of EV71 was amplified by PCR, and then was cut by restriction enzyme and inserted into expression vector pMAL-c2X. The positive recombinants were transferred into E.coli TB1, the genetically engineered bacteria including pMAL-c2X-VP2 plasmids were induced by isopropyl thiogalactoside ( IPTG) , and the expression products were analyzed by SDS-PAGE and western blotting method. EV71 IgM antibody detection method by ELISA was set up, and the sensitivity and specificity of this method was assessed; 60 neutralizing antibody positive serum samples from hand foot and mouth disease (HFMD) patients were determined, of which 52 samples were positive and 8 samples were negative; a total of 88 acute phase serum samples of HFMD patients diagnosed in clinical were also detected.</p><p><b>RESULTS</b>VP2 gene of 762 bp was obtained by PCR, the gene segment inserted into the recombinant vector was identified using restriction enzyme digestion. The recombinant vector could express a specific about 71 500 fusion protein in E.coli by SDS-PAGE. The purified recombinant protein of EV71-VP2 can react with the serum of HFMD patients to produce a specific band by western blotting. The sensitivity and specificity of ELISA was 87% and 83%, respectively. Of the 88 acute phase serum samples from children with HFMD, 48 samples (55%) were positive by the ELISA assay.</p><p><b>CONCLUSIONS</b>VP2 gene of EV71 has been cloned and a prokaryotic high expression system for VP2 gene was successfully constructed in the present study. The recombination EV71-VP2 has well antigenicity, which could be useful for developing diagnose reagent or vaccine of EV71.</p>
Texte intégral:
Disponible
Indice:
WPRIM (Pacifique occidental)
Sujet Principal:
Sang
/
Protéines recombinantes
/
Immunoglobuline M
/
Test ELISA
/
Entérovirus humain A
/
Protéines de capside
/
Allergie et immunologie
/
Anticorps neutralisants
/
Vecteurs génétiques
/
Génétique
Limites du sujet:
Humains
langue:
Chinois
Texte intégral:
Chinese Journal of Preventive Medicine
Année:
2014
Type:
Article
Documents relatifs à ce sujet
MEDLINE
...
LILACS
LIS