Immortalized Mouse Embryo Fibroblasts for DNA Repair

Isogenic SV-40 large T antigen immortalized mouse embryo fibroblasts (MEFs) that are wild type (WT), Ku70^−/−, Brca2 deficient (Brca2^m/m), Polq^−/−, and a subclone of Polq^−/-complemented by expression of human POLQ (Polq^hPOLQ) were previously described (6 (link),23 (link)). MEFs expressing DR-GFP reporter were a gift from Dr. Jeremy Stark lab (24 (link)). All MEFs were maintained in Dulbecco's modified Eagle's medium (DMEM, Corning), with 10% Bovine Calf Serum (Hyclone BCS) and 2 mM l-glutamine (ThermoFisher), and Polq^hPOLQ cells were supplemented with 2 μg/ml puromycin. Mouse ES cells (TC1) were cultured with DMEM supplemented with 15% FBS (Gibco, 1972526), 10 mM HEPES pH 7.5, 10 mM NEAA, 0.1% 2-betamercaptoethanol (Gibco, 21985023), 1% Penn-Strep (Corning, 30-002-CI) and 1:500 Leukemia inhibitory factor (LIF) treated media produced from LIF-1Cα (COS) cells. All cells were maintained at 37°C in an atmosphere of 5% CO₂. Cells in culture were routinely monitored for mycoplasma contamination using the Plasmo Test™ (Invivogen).