Cell proliferation was evaluated by MTS and EdU assays. MTS assay was performed using a CellTiter 96 AQ One Solution Cell Proliferation Assay kit (MTS, Promega, Madison, USA) according to the manufacturer’s instructions. Cells were cultured in 96-well plates at 1 × 104 cells per well. Transfected and untransfected cells were treated with 30 mg/L DADS for 24 h or left untreated, and the absorbance was recorded at 490 nm using an ELISA plate reader. Each assay was replicated 5 times.
EdU assay was conducted with an EdU kit (RiboBio, Guangzhou, China). The untreated and treated cells were incubated with the diluted EdU medium for 2 h, and then washed in PBS and subjected to DAPI staining. Finally, the images were taken and analyzed using an inverted microscope (Olympus, Tokyo, Japan). All the assays were repeated three times.
For cell cycle analysis, cells were fixed in ice-cold 70% ethanol and stained with propidium iodide (PI). The cell cycle profiles were assayed using Elite ESP flow cytometry at 488 nm and the data were analyzed using CELL Quest software (BD Biosciences, San Jose, CA, United States).
EdU assay was conducted with an EdU kit (RiboBio, Guangzhou, China). The untreated and treated cells were incubated with the diluted EdU medium for 2 h, and then washed in PBS and subjected to DAPI staining. Finally, the images were taken and analyzed using an inverted microscope (Olympus, Tokyo, Japan). All the assays were repeated three times.
For cell cycle analysis, cells were fixed in ice-cold 70% ethanol and stained with propidium iodide (PI). The cell cycle profiles were assayed using Elite ESP flow cytometry at 488 nm and the data were analyzed using CELL Quest software (BD Biosciences, San Jose, CA, United States).