Expression in Escherichia coli of cDNA fragments encoding the gene for the host-protective antigen of infectious bursal disease virus

Abstract

The larger segment of the IBDV genome codes for a 32-kDa host-protective antigen. Inserts from a cDNA library in pBR 322, containing overlapping cDNA fragments of varying sizes and covering the entire large segment of the IBDV genome, were subcloned into a mixture of expression vectors pUR 290, 291, and 292. Clones expressing the host-protective antigen, or parts of it, were identified by an immunoblot assay and the fusion proteins were further characterized by Western blot analysis using a monoclonal antibody specific for the 32-kDa polypeptide. Hybridization of inserts from expressing clones to the original cDNA library led to the identification of the region of the IBDV genome that codes for the 32-kDa host-protective antigen. Clone D1 which encodes approximately 50% and clone D6 which encodes the entire 32-kDa protein were selected for further studies. The fusion proteins from clones D1 and D6 were affinity purified and tested for their immunogenicity in chickens. Both fusion proteins induced the synthesis of antibodies in both primed and unprimed chickens that reacted specifically with denatured 32-kDa viral protein, but less well with intact virus. It was concluded that the response to the fusion proteins was to linear rather than conformational epitopes on the 32-kDa viral protein.