Development And Applacation Of Indirect ELISA For FMDV Antibody Detection And Immunological Enchancement Of New Type Adjuvants

1 Cloning and expression of vp1 gene of FMDVThe fragment of VP1 was amplified with a primer pair by RT-PCR. The interest fragment was identified by sequencing analysis. Afterwards the complete VP1 gene was amplified with an sepecific primer pair by PCR .The target gene was cloned into pMD 18-T Vector and then was subcloned into vector pGEX-4T-2.Then the recombinant was transformed into Escherichia coli BL21 (DE3) for VP1 expression. The interest gene was induced to express in E. coli with IPTG.The express product were collected at different time and subsequently were examined by SDS-PAGE and western-blotting. Results showed that the structural protein VP1 gene of FMDV can be expressed successfully in E.coli, and the molecular weight of the fusion protein was 51kD.2 Cloning and Expression of Nonstructural Protein Gene 3abc ofFMDVFoot-and-mouth disease virus(FMDV) nonstructural protein gene 3ABC was cloned into pMD18-T Vector and then was subcloned into vector pGEX-4T-2. Then the recombinant was transformed into Escherichia coli. Rosetta(DE3) for expression.. 71kD fusion protein could be tested by SDS-PAGE and can be recognized by the positive swine serum of FMDV. Through gel thin layer scanning analysis, the amount of target protein is over 15% of the total bacteria protein. The results showed that the vitro expressed protein of 3ABC via recombinant plasmid vector in the E.coli maintains antigenicity of FMDV.3 Development and application of indirect ELISA based on protein...