Plenti cmv to sv40 small large t w612 1

All plasmids were obtained from addgene (Cambridge, MA, USA): pLenti CMV/TO SV40 small + Large T (w612-1) (Addgene plasmid 22298), H2B-mCherry (Addgene plasmid 20972), Tubulin-GFP (Addgene plasmid 12298) and Centrin-2-GFP (Addgene plasmid 29559). Plk1-YFP plasmid was obtained from Dr. Leizhen Wei (University of Pittsburgh). Transfection was performed using FuGENE 6 (Roche Diagnostics, Indianapolis, IN) or lipofectamine 2000 (Life Technologies) according to the manufacture’s instructions.

Immortalized breast epithelial cell lines were cultured as described previously (18 (link)). Cells were transformed with oncogenes H-Ras^G12V, PIK3CA^H1047R, and SV40-T/t antigens expressing lentiviruses using vectors pLenti CMV-RasV12-Neo (w108-1) (HRAS G12V, #22259, Addgene), pLenti MNDU3-PGK–PIK3CA^H1047R-YFP (10 (link)), and pLenti-CMV/TO-SV40 small + Large T (w612-1) (#22298, Addgene), respectively. Cell lines in the laboratory are usually tested for Mycoplasma once in 6 months (Lonza mycoplasma testing kit, last testing was done on January 20, 2021) and cell line authentication/cross contamination yearly using marker short tandem repeat DNA sequencing method (IDEXX BioAnalytics, last testing was done on July 30, 2020). Additional details of cell culture are provided in supplementary materials and methods.