Western blot analysis was performed as described previously [12] . Briefly, a BHK-21 cell layer was infected with rWR-PRRSV-M recombinant vaccinia virus or the vaccinia virus WR strain at a multiplicity of infection (MOI) of 0.1. Cells were harvested 3 days post-infection, and total cell lysates were prepared with lysis buffer (10 mM Tris-Cl pH 7.4, 1 mM MgCl 2 , 0.5% NP40, 20 μg/ml DNase I). Cell lysates were separated by SDS-PAGE and were subsequently transferred to a membrane for Western blot analysis using an anti-PRRSV-M monoclonal antibody, HRP-conjugated goat anti-mouse IgG secondary antibody (Bio-Rad), and DAB substrate. The positive recombinant virus was named rWR-PRRSV-M.