| In this study, The recombinant piasmid pPNSl was digested with EcoR â… and Hind â…¢ to generate NSl gene, Laterly the fragment was cloned into PET30a(+) to get a prokaryotic plasmid PET-NSl; The target gene was expressed in very high level as inclusion body in the host cell BL21(DE3) when induced with IPTG The expression was optimized with proper inducing conditions of 0.6 mmol/L IPTG and 3 hours induction. The highest expression of the target protein added up to 26.7% of the total bacterial protein. Western-blot analysis proved the recombinant protein has good reactive ability against PPV positive serum. Horseradish peroxidase-staplylococcal protein (HRP-SPA) was used as the second antibody and PPA-NSl-ELISA was developed in the experiment. The optional working circumstances for the PPA-NSl-ELISA assay(antigen concentration:33ug/ml;serum dilution:1:40) with chess titration. The positive criterion of PPA-NSl-ELISA assay is OD the tested serum > 0.30. It show that detection of antibodies agaist porcine parvovirus nonstructural protein NSl may distinguish between vaccinated and infected pigs. |
