Fig. 1

BRCA2 is essential for non-transformed human mammary MCF10A cell viability. a Schematic of the BRCA2 exon3-4-floxed conditional system in MCF10A cells (filled triangle, loxP site; open circle, FRT site; Hyg, hygromycin-resistance gene). b Western blot of BRCA2 exon3-4-floxed cell extracts with or without Cre expression (asterisk, full-length BRCA2; arrowhead, ∆Ex3-4 peptide). The BRCA2 antibody Ab-1 detects BRCA2 amino acids 1651–1821. c BRCA2 ^∆Ex3-4/− cells were plated for clonogenic survival. Representative plates are shown. d Schematic of the BRCA2 ^−/− AAVS1 ^fl conditional system in MCF10A cells. Blast, Blasticidin-resistance gene. e Western blot showing BRCA2 expression in stably complemented BRCA2 ^−/− AAVS1 ^fl cells (WT, wild-type BRCA2; EV, empty vector). f BRCA2 ^−/− AAVS1 ^∆ cells were plated for clonogenic survival. g BRCA2 ^∆Ex3-4/− cells were serially passaged every 3 days. Cell number was determined at the end of each passage and normalized to the number of BRCA2 ^∆Ex3-4/+ cells at passage 0. h Cells were stained for senescence-associated β-galactosidase (SA β-gal). Left: representative images; Right: comparison of the percent SA β-gal+ cells. i Cells were quantified for apoptosis using Annexin V staining. Error bars in this figure represent one standard deviation from the mean (s.d.). n > 3. **p < 0.01; ****p < 0.0001 (unpaired two-tailed t-test)