Prokaryotic Expression And Immunological Characteristics Analysis Of The Adhesion Molecule P102-like Gene From Mycoplasma Ovipneumoniae

Mycoplasma ovipneumoniae pneumonia is a highly contagious infectious disease that can infect both sheep and goat caused by the Mycoplasma ovipneumoiae.In this study, According to the DNA sequence design of it’s gene-specific primers we extract its genome and amplified p102-like gene after the resurrection of a large number of isolates cultured MO (SZWQ). Owing to p102-like gene sequence containing a codon TGA that encodes tryptophan, TGA is the stop codon in prokaryotic expression system while the TGG encoding tryptophan. Two pairs of specific primers are designed in this research; the codon TGA of p102-like gene was changed to TGG by overlapping extension PCR method, in order to express the p102-like gene fluently in prokaryotic expression system. The results detected by SDS-PAGE shows the molecular weight of the protein are58kDa that is consistent with the expecting size of the target protein. The fusion-purified protein was detected by Western Blot detection, which shows that exogenous expression of the fusion protein has good immunoreactivity.The animal immunized testing was proceeded in mice to make purified fusion protein into vaccine. The mice were immunized four times using ELISA method to detecting whether the antibody p102-like was found in the mice sera. The results demonstrate that mice serum antibodies of p102-like were produced after the immunization; the difference between fusion protein in the experimental group and the control group was significant (P<0.05). In this immunity test, we use the humoral immune phagocytosis conditioning test and growth inhibition test to assess the effect of the fusion protein. Phagocytosis conditioning test shows that the difference between the fusion proteins the experimental group and the control group is significant (P＜0.01) and the result of growth inhibition test shows weakly positive.Through the above experiment, the trial successfully cloned and mutated the gene sequence of MO (SZWQ) of p102-like. Constructing a prokaryotic vector pET-32a-L-p102and expressing it. The various immunological experiments demonstrated that the expression of the fusion protein has some reactogenicity.