For MDA-MB-231 and BT549 cells, miRIDIAN miRNA mimics (Dharmacon, Lafayette, CO, USA) were transfected into cells using Lipofectamine 2000 (Invitrogen) according to the manufacturer’s protocol for transfecting siRNA. Mimics (final concentration of 50 nM) were mixed with 1 % Lipofectamine 2000 (v/v) diluted in Opti-MEM transfection medium (Invitrogen) and incubated for 20 minutes. The mimics were added dropwise onto cells in growth medium at a final concentration of 50 nM. Fresh medium was replaced 24 h post transfection. A non-radioactive cell proliferation assay (Promega, Madison, WI, USA) was used to assess the number of viable cells. Three biological replicates were conducted.
MDA-MB-436 and HS578T breast cancer cells were reverse transfected with 40 nM of either miR-184 mimic or scrambled using Dharmafect 4 reagent (Dharmacon) following manufacturer’s instructions; SiTox (Dharmacon) was used as a positive control. The following day, medium was changed and cells were left in culture for an additional 72 h. To assess viability, CellTiter-Glo (Promega) was added directly into the cell medium (1:2 ratio) and left to incubate for 10 minutes; luminescence was read using FLUOstar Omega plate reader (BMG Labtech, Ortenburg, Germany). The experiment was performed in three biological replicates.
MDA-MB-436 and HS578T breast cancer cells were reverse transfected with 40 nM of either miR-184 mimic or scrambled using Dharmafect 4 reagent (Dharmacon) following manufacturer’s instructions; SiTox (Dharmacon) was used as a positive control. The following day, medium was changed and cells were left in culture for an additional 72 h. To assess viability, CellTiter-Glo (Promega) was added directly into the cell medium (1:2 ratio) and left to incubate for 10 minutes; luminescence was read using FLUOstar Omega plate reader (BMG Labtech, Ortenburg, Germany). The experiment was performed in three biological replicates.