Dulbecco s modified eagle s medium

HFFs (a gift from Dr. Nicholas Wallace) were cultured in Dulbecco’s modified Eagle’s medium (Quality Biological) containing 10% fetal bovine serum (Peak Serum) and 2 mM L-glutamine (Quality Biological). Cells were incubated in a 5% CO₂ atmosphere at 37°C. VACV Western Reserve strain (ATCC VR-1354), vD9muD10mu with both decapping enzymes inactivated, and vΔA23 with intermediate transcription factor gene A23 deleted were kindly provided by Dr. Bernard Moss and were described previously (22 (link), 23 (link)). Virus titer was determined by plaque assay as described elsewhere (32 (link)).

A549 DKO, kindly provided by Dr. Bernard Moss, 293T (ATCC-CRL-3216), and HeLa (ATCC CCL-2) cells were cultured in Dulbecco’s Modified Eagles Medium (Quality Biological). BS-C-1 (ATCC CCL-26) cells were cultured in Eagles Minimal Essential Medium (Quality Biological). All growth media was supplemented with 10% FBS (Peak Serum), 2 mM L-glutamine (Quality Biological), and 100 U/mL penicillin, 100 μg/mL streptomycin (Quality Biological). Cells were grown at 37°C and 5% CO2. VACV Western Reserve (WR) strain (ATCC VR-1354) and any derived recombinant viruses (except for vD9muD10mu) were grown in HeLa cells and purified on 36% sucrose gradient and titrated by plaque assay as described elsewhere [64 (link)]. Titration of vD9muD10mu was grown and titrated in A549DKO cells as described elsewhere using anti-VACV antibody and HRP conjugated secondary antibody [25 (link), 64 (link)].