Rnase a

Trypsin-EDTA (1X), Dulbecco’s Modified Eagle Medium (DMEM), RPMI Medium 1640, and PSN Antibiotic mixture were purchased from Invitrogen (CA, USA). Fetal Bovine Serum (FBS) was purchased from Biosera (UK). Xylene cyanole and ethidium bromide (EB), Dimethylsulfoxide (DMSO) and other chemicals & reagents were purchased from Sigma Chemicals (St. Louis, USA). Sinigrin with 98% purity was purchased from Sigma-Aldrich (USA). RNase A was purchased from Amersham (USA).
ALT/SGPT (UV-Rate) & AST/SGOT (UV-Rate) Kits for detection of alanine and aspartate transferases were purchased from Stanbio Laboratory (TX, USA). Anti-GST-P polyclonal antibody (rabbit) was purchased from Medical & Biological Laboratories. Biotinylated goat anti-rabbit IgG and avidinbiotin-peroxidase complex (ABC Staining System) were purchased from Santa Cruz Biotechnology (CA, USA).

The cells were treated as indicated. After 48 h treatment, 4 × 10⁶ cells were washed twice with cold PBS containing 1% FBS. After ethanol fixation (at least 12 h at 4°C), cells were washed twice with cold PBS. The pelleted cells were stained by adding PBS containing PI (Sigma-Aldrich) and RNase A (Amersham). Staining was achieved in the dark at 4°C for 3 h prior to flow cytometry analysis using a BD LSR Fortessa flow cytometer, and analyzed with the Tree Star FlowJo (Ashland, OR, USA) software program to model the cell cycle assays. Fluorescence data were collected using a 561 nm laser excitation, and emission was collected using a 610/20 filter. Fluorescence data were obtained from at least 10,000 viable cells per sample.

MDA‐MB‐231 cells were washed twice with cold PBS, scraped, and collected by centrifugation at 1500 × g for 5 min. Nuclear and cytoplasmic proteins were extracted using the Nuclear‐Cytosol Extraction Kit (Applygen Technologies, Beijing, China). The supernatant (nuclear fraction) was collected by centrifugation at 13 000 × g for 30 min at 4 °C. The protein concentration was determined using the BCA Pierce protein assay kit (Thermo Fisher Scientific) and MDA‐MB‐231 nuclear proteins (6 mg) were used for the FPLC assay. The proteins were concentrated to 1 mL using a Millipore Ultrafree centrifugal filter apparatus (10 kDa nominal molecular mass limit) and then applied to an 850 × 20 mm Superose 6 size exclusion column (Amersham Biosciences) that was equilibrated with buffer D (20 × 10^‐3m HEPES, pH 8.0, 10% glycerol, 0.1 × 10^‐3m EDTA, and 300 × 10^‐3m NaCl) containing 1 × 10^‐3m dithiothreitol and calibrated with protein standards (blue dextran, 2000 kDa; thyroglobulin, 669 kDa; ferritin, 440 kDa; catalase, 232 kDa; bovine serum albumin, 67 kDa; and RNase A, 13.7 kDa; Amersham Biosciences). The column was eluted at a flow rate of 0.5 mL min^‐1 and fractions were collected.