Establishment and Characterization of Notch3-Overexpressing Ovarian Cancer Cell Lines

The mouse ID8 IP2 cell line was developed in the lab of co-author Jill Slack-Davis and provided directly to the Kitajewski lab [29 (link)]. ID8 IP2 cells were cultured in DMEM with 10% FBS, 1% ITS Liquid Media Supplement (Sigma-Aldrich I3146), and 1% penicillin-streptomycin, and infected with lentiviral vector FUW-luciferase-mCherry [71 (link)]. Human ovarian cancer cell lines OVCAR3. OVSAHO, and OVCA429 were a kind gift of Dr. Joanna Burdette (University of Illinois Chicago), A2780 was a kind gift of Dr. Tian-Li Wang (Johns Hopkins), and SKOV3-IP1 was a kind gift of Dr. Olga Razorenova (University of California Irvine). OVCAR3, A2780, OVSAHO and SKOV3-IP1 were cultured in RPMI 1640 with 10% FBS and 1% penicillin-streptomycin, while OVCA429 was cultured in MEM with 10%FBS, 1% L-glutamine, 1% NEAA, 1% Sodium pyruvate and 1% penicillin-streptomycin. Mouse and human cells were lentivirally infected with virus derived from a pCCL vector encoding an HA-tagged Notch3 intracellular domain (codons 1664–2318) followed by an IRES-GFP (Notch3IC, Fig 1C) or empty pCCL vector. Five matched sets of Notch3IC and control ID8 IP2 lines were generated, identified as Sets #1-#5 throughout. Human cells were freshly infected prior to each experiment. Post-hoc testing suggests that all ID8 IP2 lines were mycoplasma positive throughout these experiments.