Culturing and Transfecting Cell Lines

The previously described F9 and dyskerin-mutated F9 cells (F9-A353V) [19 (link), 21 (link)], were cultured in Dulbecco Modified Eagle Medium (DMEM) supplemented with 10% foetal bovine serum, 2 mM glutamine (Gibco) and 1.5 gr/ml of sodium bicarbonate. HEK293T cells (ATCC) were cultured in DMEM media supplemented with 10% foetal bovine serum and 2 mM glutamine. DC-C and DC-3 cells (Coriell Cell Repository) were cultured in Roswell Park Memorial Institute (RPMI) media supplemented with 20% foetal bovine serum and 2 mM glutamine. XDC-1787-C (Corriel Cell Repository) and F26IIB cells (generated in our laboratory), were cultured in Minimum Essential Medium Eagle (MEM) media supplemented with 15% foetal bovine serum and 2 mM glutamine. Cells were transfected with 10 μg of DNA/10⁶ cells using lipofectamine plus (Invitrogene, Carlsbad, USA) or the K2 transfection kit (Biontex Lab, Planegg/Martinsried, GE) according to the manufacturers’ instructions. 15 μg of peptides per 35 mm dish were transfected using the ProteoJuice Protein Transfection Reagent (Merck Millipore, Billerica, MA, USA) or the K2 transfection kit.