Mda mb 157 cells

MDA-MB-157 cells (ATCC, Manassas, VA) and HEK293T cells were cultured in DMEM supplemented with 10% fetal bovine serum, Penicillin and Streptomycin and GlutaMax (Thermo Fisher Scientific). Viral barcoding vectors were transfected into subconfluent HEK293T cells with pCL-10A1 retroviral packaging plasmid using Fugene 6 (Promega, Madison, WI) and viral supernatant was collected and concentrated with polyethylene glycol (PEG). For concentration, viral supernatant was spun at 931 gravities for 4 min and decanted into 1/5.5 volumes of sterile 50% PEG-3350 in phosphate-buffered saline (PBS). After an overnight 4° incubation, the mixture was spun for 1455 gravities for 20 min, decanted, spun again at 524 gravities for 4 min, and the pellet resuspended in PBS with 1% bovine serum albumin and frozen at −80°C. For infection, 1 × 10⁴ cells were incubated with concentrated virus and 30 µg ml⁻¹ protamine sulphate and spun at 1455 gravities for 1.5 h. Viral concentration was optimized to infect approximately 10% of cells. After 48 h, cells were selected with 3 µg ml⁻¹ puromycin to kill uninfected cells. Barcoded cells were expanded without discarding cells until the population was at least 2 × 10⁷ cells, and subsequently split into subpopulations no smaller than 2 × 10⁶ cells to maintain clonal representation.