Expression and purification of an adhesive protein of rabbit Pasteurella multocida C51-3 and detection of its antigenicity.
- Author:
Wulumuhan NAZIERBIEKE
1
;
Fang YAN
;
Cui HE
;
Lei ZHANG
;
Entomack BORRATHYBAY
Author Information
1. Laboratory of Bioengineering, College of Biology and Environmental Sciences, Jishou University, Jishou 41600, China.
- Publication Type:Journal Article
- MeSH:
Adhesins, Bacterial;
biosynthesis;
genetics;
immunology;
Animals;
Cloning, Molecular;
Escherichia coli;
genetics;
metabolism;
Pasteurella multocida;
chemistry;
Rabbits;
microbiology;
Recombinant Fusion Proteins;
biosynthesis;
genetics;
immunology
- From:
Chinese Journal of Biotechnology
2008;24(8):1446-1453
- CountryChina
- Language:Chinese
-
Abstract:
The cp36 gene encoding an adhesive protein was amplified by PCR from genomic DNA of rabbit P. multocida C51-3 strain, and cloned into the pMD18-T vector and then sequenced. The mature adhesive protein without a signal peptide of cpm36 gene was amplified by PCR from the recombinant plasmid pMD18-cp36, then cloned into the prokaryotic expression vector pQE30 to provide a recombinant plasmid pQE30-cpm36. The recombinant protein of CPM36 was produced in Escherichia coli M15 harboring the recombinant plasmid pQE30-cpm36 by IPTG induction, and the recombinant protein purified by the affinity chromatography with Ni(2+)-NTA resin. The sequence analyses showed that the ORF of cp36 gene was 1032 bp in length, and DNA homology of the cp36 genes between the C51-3 strain and the previously reported different serotype strains of P. multocida in GenBank was 76.9 to 100%. The SDS-PAGE analyses revealed a single fusion protein band with a molecular weight of 37 kD, and the Western blotting analysis demonstrated that the recombinant protein CPM36 and native 36 kD protein of C51-3 were recognized specifically by an antiserum against the recombinant protein, suggesting that the recombinant protein is an antigenic protein.
