Cloning and expression of p25 protein gene(p25) of Riemerella anatipestifer serotype 1 and its detection of immunogenicity

In order to study the immunogenicity of p25 protein of Riemerella anatipestifer serotype 1,the whole ORF of p25 gene was amplified from RA serotype 1 genomic DNA.The bioinformatics indicated that p25 gene was an unknown function conserved domain contianing an complete CDD named PRK00110.After the amplified product was digested by endonuclease BamH Ⅰand Hind Ⅲ,the p25 gene was cloned into prokaryotic expression vector pET-32a(+) and the recombinant expression vector pET-32a(+)-p25 was constructed.The recombinant expression vector pET-32a(+)-p25,with the correct sequence,was introduced into E.coli Rosetta and expressed successfully in E.coli Rosetta.SDS-PAGE analysis showed that the MW of recombinant protein was about 45 000 and the recombinant protein was mainly existed as soluble protein.Western blotting indicated that this recombinant p25 protein had good immunogenicity.