4.5. Cell Culture and Transfection

MCF7 cell line was cultured in high glucose Dulbecco’s modified Eagle’s medium (DMEM) supplemented with 10% fetal bovine serum (FBS) and 1% penicillin/streptomycin. MCF12A cells were cultivated in a special medium containing DMEM/F12 (Thermo Fisher Scientific, Waltham, MA, USA), 5% heat inactivated horse serum (Gibco, Grand Island, NY, USA), 20 ng/mL recombinant human EGF (AF100-15 Peprotech, Rocky Hill, NJ, USA), 0.5 µg/mL hydrocortisone, 100 ng/mL cholera toxin (C-8052, Sigma, St. Louis, MO, USA), 10 µg/mL human recombinant insulin (Zinc solution 12585-014, Gibco, TN, USA), and 1% penicillin/streptomycin and incubated at 37 °C with 5% humified CO₂. To examine the promoter activity of the cloned regions, the recombinant vectors were used to transfect MCF7 cells. All experiments were done with complete medium in at least triplicates.