HCT 116p53−/− cells and rpL3ΔHCT 116p53−/− cells, derived from HCT 116p53−/− cell line and stably silenced for rpL3, were cultured in Dulbecco's Modified Eagle's Medium (DMEM) with glutamax (Invitrogen, Carlsbad, California) supplemented with 10% fetal bovine serum (FBS), 2 mM L-glutamine, penicillin-streptomycin 50 U/ml and 0.5 μg/ml puromycin (Sigma-Aldrich).
CBSΔHCT 116p53−/− cell line was obtained from HCT 116p53−/− cells as previously reported [34 (link)]. Cells were transfected with 2 μg of different shRNA CBS plasmids (Sigma-Aldrich) by using Lipofectamin 2000 (Life Technologies) according to the manufacturer's instructions. Stable clones were selected in medium containing 1 mg/ml of Puromicin (Sigma-Aldrich) and assayed for the detection of CBS expression level by western blotting.
shRNA transfections were performed in cells as previously described [34 (link)].
Drug treatments were performed by adding to cells 100 μM 5-FU (Sigma-Aldrich, St. Louis, MO, USA), CHX
CBSΔHCT 116p53−/− cell line was obtained from HCT 116p53−/− cells as previously reported [34 (link)]. Cells were transfected with 2 μg of different shRNA CBS plasmids (Sigma-Aldrich) by using Lipofectamin 2000 (Life Technologies) according to the manufacturer's instructions. Stable clones were selected in medium containing 1 mg/ml of Puromicin (Sigma-Aldrich) and assayed for the detection of CBS expression level by western blotting.
shRNA transfections were performed in cells as previously described [34 (link)].
Drug treatments were performed by adding to cells 100 μM 5-FU (Sigma-Aldrich, St. Louis, MO, USA), CHX
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