Development Of Subunit Vaccine With Mycoplasma Bovis P81 Protein Expressed In E.coli

Mycoplasma bovis belongs to Mollicutes,Mycoplasmatales,Mycoplasma genus,less envelope.The size is 0.2-0.3 ?m,Mycoplasma bovis isolated from the milk of affected mastitis in America,1961.Mycoplasma bovis isclosely related to respiratory disease,1976.Mycoplasma bovis first isolated in China,2008.Mycoplasma bovis causes pneumonia?mastitis?arthritis?keratoconjunctivitis,et al,which has become major pathogenic bacteria and obstructs the development of breed.Mycoplasma bovis has gradually spread in China,and causes large economic loss.Research and development of subunit vaccine has become an important task at present.Cellulose has infinite extensibility.Cellulose is linked to several specific proteins,respectively.Prevent aggregation during self-assembly or expression.Then,they induced to assemble together at a specific time and place.Cellulose,as a new immune adjuvant,has good immune enhancement effect,and provide new possibilities for the development of multiple vaccine.This study selected P48 protein and P81 protein.Amplification of 16SrRNA,P48 and P81 genes of Mycoplasma bovis,neighbour-joining tree analysis the sequencing results.Overlapextension PCR can mutate the synonymous codon TGA to TGG.Insert the target fragment into the pET-32a(+),Transformation recombinant plasmid into competent cells BL21(DE3)in E.coil.Analysis thereactiono-genicity of recombination protein by Western blot.The indirect ELISA method of P81 protein was established successfully;The P81 animal testing have an antibody potency reached 212 in 21d.