HEK293 Cell Culture and Transfection

Low-passage-number HEK293 cells (ATCC, Manassas, VA, USA) were maintained in DMEM supplemented with 10% FBS and 100 μg ml⁻¹ penicillin-streptomycin at 37°C. In all experiments, cells were transiently transfected with 1 μg T-antigen and a total of 6 μg of the respective DNA plasmids, using the calcium phosphate precipitation method. For flow cytometry assays, hERG channel subunits were transfected in 1:1 (or 3:3 μg, BBS-hERG 1a-YFP + 1b-CFP or hERG 1a-YFP + 1b-CFP) ratio. Cells were cultured in supplemented DMEM at 37°C for 24 h before transfection.