Eukaryote Expression Of The ORF2 Gene Of Porcine Circovirus Type 2 And Development Of ELISA Based On Recombinant Protein

ORF2 gene is a major protein of porcine circovurus type 2, which has nature immunity . From recombinant plasmid pMDORF2 by using enzyme, ORF2 gene was cutted and ligated with vector pEGFP-Cl then transformed it into DH5a. Cutted off frame shift base by using Bgl â…¡ and Hindâ…¢then filled in it with Klenow , transformed it into DH5a. Transfected recombinant plasmid pEGFP-Cl-ORF2 into IBRS-2 cell to express, the expressed recombinant protein could be discerned by PCV-2 positive serum and had favorurable antigenicity based on Western-blot detection. Took the recombinant fusion protein as antigen, established indirect ELISA method for dectecting PCV-2 antibody. To validate the assays, ORF2 ELISA was performed with 48 serum samples collected from different herds in the Sichuan. Compared with IDEXX, the diagnostic sensitivity, specificity, and accuracy of 0RF2 ELISA were similar (>90%). The tests showed no cross-reactivity with antibodies to PPV, PRV, PRRSV, HCV. Modified indirect reaction condition of ELISA: the antigenic optimal coating concentration 3. 6ug/ml, the optimal detection serum dilute strength 1:40.