Secretive Expression and identification of vp2, the Major Antigenic Protein of Infectious Bursal Disease Virus in Yeast
Abstract: Here we report that the major antigenic protein VP2 of very virulent Infectious bursal disease virus (vvIBDV) was secretively expressed with high level in yeast (Pichia pastoris) expression system. Primers were designed to amplify the previously cloned vvIBDV-vp2 gene from the plasmid pMD18-T- VP2. The amplified 1.4kb vp2 gene fragment was sub-cloned into the yeast-expressing plasmid pPICZα-A. The recombinant pPICZα-A-VP2 was analyzed by restriction enzymes and sequenced, which was then linearized by SacⅠand transfected into Pichia pastoris X-33. After selection with ZeocinTM, the phenotype identity, the inductive expression, the SDS-PAGE and the western blot analysis of culture supernatant, an engineering Pichia pastoris strain X-33/pPICZαA-VP2 with which the VP2 protein could be expressed with high level was obtained. SDS-PAGE and Western blot analysis indicated that the expressed product of the VP2 in culture supernatant of X-33/pPICZαA-VP2 was about 55kDa, a bit larger than expected. Gel scanning and Bradford protein analysis showed that the expressed product was as high as 23mg/ L, or 28% of total proteins in culture supernatant of X-33/ pPICZαA-VP2.