Expression of hepatitis B core antigen gene in recombinant vaccinia virus

Plasmid pMM2066 was digested with EcoRI restriction enzyme and an 870 bp fragment separated; it was located 12 bp upstream from the ATG codon of the antigen C structural gene. This DNA fragment was then inserted into shuttle vector pGJP-5 just downstream from the P7.5 promoter, producing recombinant vector p5-C2066. Three TK- recombinant vaccinia viruses expressing HBcAg were isolated by plaque assay, using homologous recombination in TK- 143 cells in the presence of BUdR. The supernatant from cells infected with recombinant virus (in dilutions of up to 1:64) still gave significant positive ELISA results. HBeAg activity titer was about the same. The HBcAg particles (26.6 nm) were visualized by electron microscopy. The influence of infection multiplicity, cell type, and culturing temperature on HBcAg expression was also investigated.