Bacterial expression of an immunologically reactive PCV2ORF2 fusion protein

The entire coding region of open reading frame 2 (ORF2) of porcine circovir us 2 (PCV2) was: linked to the 3'-end of the maltose-binding protein (MBP)- His(8)-tag gene. The fusion protein was expressed as soluble form after ind uction by isopropylthio-beta -D-galactoside. MBP-His(8)-ORF2 was purified t o homogeneity by immobilized metal affinity chromatography based on the int eraction of the polyhistidine-tag with metal ions. Expression could represe nt 1% of the total protein in Escherichia coli, allowing approximately 1 mg of highly purified protein to be obtained per liter of bacterial culture. The fusion protein was recognized in Western blot by anti-PCV2 polyclonal a ntibody and swine sera with PCV2 infection. In addition, rabbit polyclonal antibody raised against purified MBP-His,ORF2 fusion protein reacted with t he ORF2 protein in immunoprecipitation, The availability of this fusion pro tein should permit a thorough study of prevalence of PCV2 infection in larg e-scale serological studies of field samples. (C) 2001 Academic Press.