APC Knockdown in Breast Cancer Cells

MMTV-PyMT;Apc^+/+ and MMTV-PyMT;Apc^Min/+ cells were isolated as previously described [23 (link)] and were grown in RPMI 1640 media supplemented with 10 % fetal bovine serum, 1 % penicillin/streptomycin and 1:5000 plasmocin (Invivogen, San Diego, CA). MDA-MB-157 breast cancer cells (ATCC, Manassas, VA) were maintained in RPMI 1640 media supplemented with 10 % fetal bovine serum, 1 % penicillin/streptomycin, 25 mM HEPES and 1:5000 plasmocin. All cells were routinely passaged using 0.25 % trypsin/EDTA and maintained at 37 °C with 5 % CO₂. MDA-MB-157 cells were subjected to lentiviral mediated shRNA knockdown of APC using two different MISSION shRNA APC constructs (Sigma-Aldrich, St Louis, MO). After transduction, cells were maintained in media containing 1.5 μg/mL puromycin (Sigma-Aldrich).