The ESCC cell lines were treated with 1.5 mmol/L 5-Aza-20-deoxycytidine (Sigma A3656) for 96 h. Twenty-four hours before harvest, 0.5 mmol/L trichostatin A (Sigma T8552) was added. DNA, RNA, and protein were extracted and analyzed for the methylation status of the miR-10b-3p promoter as well as the expression of miR-10b-3p and its target proteins.
5 aza 20 deoxycytidine
Manufactured by Merck Group
Sourced in United States
5-Aza-20-deoxycytidine is a nucleoside analog that inhibits DNA methyltransferase activity. It is used as a research tool in molecular biology and epigenetics studies.
Automatically generated - may contain errors
Lab products found in correlation
Fetal bovine serum (fbs), by Thermo Fisher Scientific (2 mentions)
Trichostatin a, by Merck Group (1 mentions)
Astrocyte media, by ScienCell (1 mentions)
Fetal bovine serum (fbs), by ScienCell (1 mentions)
Dld 1, by Japanese Collection of Research Bioresources Cell Bank (1 mentions)
Fetal bovine serum (fbs), by Lonza (1 mentions)
Rpmi 1640, by Nissui Pharmaceutical (1 mentions)
Rabbit anti il 6, by Bioworld Technology (1 mentions)
Bpv pten, by Merck Group (1 mentions)
Rabbit anti pten, by Abcam (1 mentions)
Rabbit anti dnmt1, by Cell Signaling Technology (1 mentions)
Complete freund s adjuvant, by Chondrex (1 mentions)
Interleukin 6 (il 6), by Sangon (1 mentions)
Ccl 2, by Sangon (1 mentions)
Tnf α, by Sangon (1 mentions)
Ccl 3, by Sangon (1 mentions)
Vcam 1, by Sangon (1 mentions)
Decitabine, by Merck Group (1 mentions)
5 protocols using 5 aza 20 deoxycytidine
1
Epigenetic Regulation of miR-10b-3p
2
Exploring miR-98 Expression in Glioma Cells
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Normal human astrocytes were purchased from the IBS Cell Bank of Fudan University (Shanghai, China) and cultured in astrocyte media (Science Cell, Carlsbad, CA, USA) with 10% fetal bovine serum (FBS) at 37°C in a humidified incubator containing 5% CO2. Human glioma cell lines, including U87, U251, U373 and SHG44, were purchased from the Cell Bank of Central South University. Cells were cultured in Dulbecco's modified Eagle's medium (DMEM) with 10% FBS (both from Thermo Fisher Scientific, Waltham, MA, USA) at 37°C in a humidified incubator containing 5% CO2. 5-Aza-20-deoxycytidine (5-Aza) was purchased from Sigma-Aldrich; Merck KGaA (St. Louis, MO, USA), and dissolved in phosphate-buffered saline (PBS) at indicated concentrations. Glioma cells were treated with 1 mM 5-Aza for 48 h, followed by assessment of miR-98 expression.
3
Cell Line Demethylation Assay
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Four GC cell lines of the MKN series and CRC cell line (DLD-1) were purchased from the Japanese Collection of Research Bioresources Cell Bank (Osaka, Japan). All cell lines were maintained in RPMI-1640 (Nissui Pharmaceutical, Tokyo, Japan) containing 10% fetal bovine serum (BioWhittaker, Walkersville, MD, USA), 2 mM L-glu- tamine, 50 U/mL penicillin, and 50 lg/mL streptomycin in a humidified atmosphere of 5% CO 2 and 95% air at 37 °C. Cells were treated with 1 lM 5-aza-2 0 -deoxycytidine (Sigma Chemical, St Louis, MO, USA) for 48 h to achieve demethylation.
4
Investigating PTEN Regulation and Epigenetics
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Bpv (PTEN) and 5-Aza-2 0 -deoxycytidine were obtained from Sigma Inc. (St. Louis, MO, USA). Complete Freund's adjuvant was purchased from Chondrex, Inc. (WA, USA). Rabbit anti-PTEN and anti-TNF-a monoclonal antibody were obtained from Abcam (Cambridge, UK). Rabbit anti-DNMT1 and anti-Vimentin (Alexa Fluor 594 Conjugated) were acquired from Cell Signaling Technology (Danvers, MA, USA). Rabbit anti-IL-6 and anti-b-actin monoclonal antibody were purchased from Bioworld (Shanghai, China). Peroxidase-conjugated goat antirabbit IgG (H þ L) was obtained from ZSGB-BIO (Beijing, China). PTEN, TNF-a, IL-6, IL-1b, CCL-2, CCL-3, VCAM-1, and VEGF-a and b-actin primers were obtained from Sangon Biotech (Shanghai) Co., Ltd (Shanghai, China).
5
Decitabine Regulates WIF-1 Expression
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The expression of the Wnt pathway inhibitor WIF-1 is related to the methylation of its promoter. Because decitabine, which is also called 5-aza-2 0 -deoxycytidine (Sigma-Aldrich, St. Louis, MO), could regulate the methylation of the promoter and promote the expression of WIF-1. Log-phase MMQ and GH3 cells (.99% viable) were plated into six-well culture plates (;1310 6 cells/well) and were treated with decitabine solution (10 mM) combined with the miR-137 mimics or miR-137 mimics control (100 nM). After treatment of 72 hours, the cells were harvested for the MTS assays and protein assays.
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