In vivo minigenome assay of RSV.

BSR-T7 cell monolayers grown on 6-well plates were infected with vTF7-3 at 37°C for 1 h. The plasmids pGBDT7-N (1.0 μg), pGADT7-P (1.0 μg), pBS-L (0.25 μg), pBS-minigenome-CAT (1.0 μg), and pGBDT7-M2-1 (1.5 μg), encoding N, P, L, M2-1, and RSV minigenome, respectively, were transfected with Lipofectamine 2000 (Invitrogen). At 5 h posttransfection, the transfection medium was replaced by DMEM containing 4% FBS. At 24 h posttransfection, the medium was replaced by DMEM containing 4% FBS with 0.25, 0.5, 1.0, or 2.0 μM TSA or mock treated. At 48 h posttransfection, cells were harvested, and the cell lysates were subjected to a chloramphenicol acetyltransferase (CAT) enzyme-linked immunosorbent assay (ELISA) for detection of the relative CAT activity as described by Chen et al. (30). All assays were repeated at least three times for accuracy.