Expression Profiling of HEK293T/c17 Cells

HEK293T/c17 cells, which do no not express detectible DPD (21 (link)), were acquired from the American Type Culture Collection and cultured as previously detailed (21 (link)). Low passage stocks were prepared after receipt and cells were maintained in culture media for a maximum of 10 total passages after receipt. The cells were regularly monitored by microscopy to confirm cell identity by monitoring cell morphology. Mycoplasm screening was performed every 3 months or whenever a question of contamination arose by using the mycoplasma detection kit PlasmoTest (InvivoGen, San Diego, CA). Additionally, cell proliferation was periodically monitored using real-time cell analysis (xCelligence RTCA System, Acea Biosciences, San Diego, CA). For transfection, 1×10⁶ cells were seeded in 6-well plates containing Dulbecco's Modified Eagle's Medium (Mediatech, Manassas, VA) supplemented with 10% Fetal Bovine Serum (FBS) (Denville Scientific, Holliston, MA) and 1% L. Glutamine, (Thermo Scientific, Waltham, MA) at 37° C for 24 hours prior to transfection. Cells were transfected using XtremeGENE HP (Roche Diagnostics, Indianapolis, IN) per manufacturers protocol. Media was replaced after 24 hours, and protein lysates were prepared 48 hours after transfection as described previously (21 (link)).