Cell lines and culture conditions

Breast cancer cell lines MCF-7, T47D, ZR-75-1, BT474, HCC1500, HCC1937, MDA-MB-361, MDA-MB-231, MCF10A normal human mammary epithelial cells, HEK293T human embryonic kidney cells, and HeLa human cervical adenocarcinoma cells were obtained from ATCC along with authentication, and cultured according to the supplier’s instructions. Cell lines maintained for more than 20 passages were further authenticated using short tandem repeat genotyping. All cells were routinely monitored for mycoplasma infection using a Mycoplasma Detection Set (TaKaRa, Japan). Cells stably expressing BRCA1-specific short hairpin RNA (shRNA) in doxycycline (Dox)-inducible manner were established by lentiviral infection with CS-RfA-ETBsd comprising targeting sequences, followed by selection with blasticidin, as previously described [15]. The cells were treated with 1 µg/ml Dox for 48 h prior to experimentation. Tamoxifen-resistant cell lines were established by culturing the cells in a medium containing IC₉₀ doses of 4-hydroxytamoxifen (4-OHT) for 24 h, washed, and further cultured in a medium containing IC₅₀ doses of 4-OHT for approximately one month for MCF-7 and 3 months for T47D and ZR-75-1 cells, until cells recovered to exponential growth states. Cells were then maintained with 5 µM 4-OHT unless otherwise described. Cells treated with ionizing irradiation (IR) were exposed to the indicated doses of X-ray irradiation and cultured for the stated times before analysis. The chemical agents that were used are listed in Table ​Table1.1. The concentration of the agents was determined by preliminary examinations based on the manufacturer's instructions and on their effects on appropriate markers.

Chemical agents used in the study