Hct116 p53wt

HCT116 p53 WT and p53 KO (Colon cancer) cell lines were purchased from American Type Culture Collection (Manassas, VA, USA). Cells were cultured in Dulbecco’s modified Eagle’s medium (DMEM; Gibco) plus 10% fetal bovine serum (FBS; Gibco) supplemented with 1% penicillin-streptomycin (Gibco) at 37 °C and 5% CO2. To generate stable transfection cell lines, SENP3 (WT) and SENP3 (9A) was cloned into the lentivector pCDH (System Biosciences, #CD510B-1). Production of Lentivirus and infection were carried out according to user manual as previously reported^{32 (link)}. The cells were selected with 2 μg/mL puromycin (Amresco) for 1 week to get SENP3 (WT) and SENP3 (9A) stably transfected cells.

HEK-293T cell and human colon cancer cell line HCT116 (p53 WT) were obtained from the American Type Culture Collection (ATCC). HCT116 (p53 Null) cell was generated by using CRISPR/Cas9, and donated as a kind gift from Dr. Dan Luo 13 (link). HEK-293T cells were maintained in Dulbecco's modified Eagle's medium (DMEM), supplemented with 10% fetal bovine serum (PAN, Germany), 100 U/mL penicillin and 100 µg/mL streptomycin (GIBCO, Rockville, MD, USA). HCT116 cells were cultured in McCoy's 5A medium (Hyclone Inc, USA), supplemented with 10% fetal bovine serum (PAN, Germany), 100 U/mL penicillin and 100 µg/mL streptomycin (GIBCO, Rockville, MD, USA). All cells were maintained at 37 °C in a humidified incubator under 5% CO₂.
Lentiviral particles were prepared as previously study 14 (link). For the generation of stable cell lines, cells were infected with recombinant lentiviruses, then treated with 2 μg/mL puromycin (Sigma Inc, USA) at 48 h of post-infection for generation of stable cells expressing desired gene or shRNA.