Mcf 7 human breast tumor cells

MCF-7 human breast tumor cells were obtained from the American Type Culture Collection (Manassas, VA, USA). MCF-7 cells were cultured in Dulbecco’s Modified Eagle’s Medium (Corning Inc, Tewksbury, MA, USA) with 10% fetal bovine serum (Corning Inc) at 37°C in 5% CO₂. The Lipofectamine 2000 transfection kit (Invitrogen, Carlsbad, CA, USA) was used according to the manufacturer’s instructions. Human AKT1-specific siRNA (siAKT1) and control siRNA (siNC) were supplied by Shanghai GenePharma Co, Ltd (Shanghai, People’s Republic of China) as follows: AKT1 sense, 5′-UGCCCUUCUACAACCAGGATT-3′; AKT1 antisense, 5′-UCCUGGUUGUAGAAGGGCATT-3′; control sense, 5′-UCCGUUUCGGUCCACAUUCTT-3′; and control antisense, 5′-GAAUGUGGACCGAAACGGATT-3′. Fluorescein-tagged siRNA (FAM-siRNA) was synthesized by modifying the 3′-end of the AKT1 siRNA sense strand. Mouse anti-AKT1 and anti-glyceraldehyde-3-phosphate dehydrogenase were purchased from Abcam (Cambridge, UK).