Geneprint kit

MCL cell lines Mino (CRL-3000), JeKo-1 (CRL-3006) and REC-1 (CRL-3004), were obtained from American Type Culture Collection (ATCC). To avoid Mycoplasma contamination, cell lines were routinely tested for Mycoplasma infection by PCR. The identity of all cell lines was verified by using GenePrint® kit (Promega, Madison, WI, USA). MCL cell lines were cultured in RPMI 1640 complemented with 10–20% fetal bovine serum (FBS), 2 mM L-glutamine and 50 μg/mL penicillin/streptomycin (Life Technologies, Carlsbad, CA, USA) and grown in a humidified atmosphere at 37 °C with 5% CO₂. The murine bone-marrow derived stromal cell line OP9 (CRL-2749; ATCC) overexpressing DLL4 (OP9-DLL4) was generated and grown as described [17 (link), 18 (link)]. Primary cells from MCL patients were isolated and cultured as described [19 (link)] and conserved within the Hematopathology collection of our institution registered at the Biobank from Hospital Clínic-IDIBAPS (R121004–094). The ethical approval for this project including informed patient consent was granted following guidelines of the Hospital Clínic Ethics Committee.

COV318 cells (Public Health England) were cultured in DMEM (Sigma-Aldrich or Gibco, Thermo Fisher Scientific) supplemented with 10 % fetal bovine serum (FBS, Gibco, Thermo Fisher Scientific), 100 U/ml penicillin, 100 µg/ml streptomycin (all from Sigma-Aldrich) and 2 mM glutamine (Gibco). KURAMOCHI (Tebu-Bio) and OVCAR4 (kindly provided by Prof. Charley Gourley, University of Edinburgh) and OVCAR8 (kindly provided by Prof. Kaisa Lehti, University of Helsinki) cells were cultured in RPMI (Gibco) supplemented with 10 % FBS, 2 mM glutamine (Gibco), 100 U/ml penicillin, 100 µg/ml streptomycin (all from Sigma-Aldrich). Cells were cultured at 37 °C, 5 % CO₂ and tested negative for mycoplasma. Cell lines were authenticated using gDNA extracted from the cells using the Puregene Gentra Kit and multiplexed using the Promega Geneprint Kit. Samples were run on an Applied Biosystems 3130xl DNA analyser and analysed using Applied Biosystems Genemapper v4.1 software. Profiles were compared with ATCC (LGC standards) and both Cellosaurus and DSMZ databases.

The human cell lines HEK293-T cells (gift from Dr. Mounira Amor-Gueret) were cultured in DMEM (Eurobio Abcys, Courtaboeuf, France) media containing 25 mM sodium bicarbonate and 2 mM l-glutamine supplemented with 10% FBS (EuroBio Abcys). The BRCA2-deficient colorectal adenocarcinoma cell line DLD1 BRCA2^{[−/−31 (link)} (HD 105-007) and the parental cell line DLD1 BRCA2^+/+ (HD-PAR-008) were purchased from Horizon Discovery (Cambridge, England). The cells were cultured in RPMI media (EuroBio Abcys) containing 25 mM sodium bicarbonate and 2 mM l-glutamine (EuroBio Abcys) Supplemented with 10% FBS (EuroBio Abcys). The DLD1 BRCA2^−/− cells were maintained in growth media containing 0.1 mg/ml hygromycin B (Thermo Fisher Scientific). The stable cell lines of DLD1^−/− BRCA2-deficient cells expressing BRCA2 WT or variants of interest generated in this study were cultured in growth media containing 0.1 mg/ml hygromycin B (Thermo Fisher Scientific) and 1 mg/ml G418 (Sigma-Aldrich).
All cells were cultured at 37 °C with 5% CO₂ in a humidified incubator and all cell lines used in this study have been regularly tested for mycoplasma contamination (MycoAlert, Lonza) and genotyped using GenePrint kit (Promega).