5 aza

Manufactured by Thermo Fisher Scientific

Sourced in China

5-Aza is a chemical compound used in laboratory research. It is an analog of the nucleoside cytidine, with the carbon-5 position modified to contain a nitrogen atom instead of a carbon atom. This structural change confers unique properties that make 5-Aza useful for specific experimental applications.

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Lab products found in correlation

5 aza, by Merck Group (4 mentions) Rpmi 1640 medium, by Thermo Fisher Scientific (2 mentions) Fetal bovine serum (fbs), by Thermo Fisher Scientific (1 mentions) Lip2000, by Thermo Fisher Scientific (1 mentions) Cell counting kit 8 assay, by Dojindo Laboratories (1 mentions) Elisa reader, by Bio-Rad (1 mentions) Trypsin edta, by Thermo Fisher Scientific (1 mentions) 5 aza 2 deoxycytidine 5 aza, by Merck Group (1 mentions) Dimethyl sulfoxide (dmso), by Merck Group (1 mentions) Mcf 7, by American Type Culture Collection (1 mentions) Fetal bovine serum (fbs), by Capricorn (1 mentions)

4 protocols using 5 aza

Cell Line-Based Epigenetic Regulation Study

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HBE, A549, H460, and H1299 cell lines, kindly provided by Cell Bank, Chinese Academy of Sciences (Shanghai, People’s Republic of China), were maintained in RPMI-1640 medium (Thermo Fisher Scientific, Waltham, MA, USA) with 10% fetal bovine serum (Thermo Fisher Scientific) in humidified 5% CO₂ at 37°C.
The cells were cultured in 6-well plates (5×10⁵ cells/well) for 24 h and then exposed to 5 μmol/L 5-Aza (Sigma, St Louis, MO, USA) for 72 h. The cells were collected for quantitative real-time polymerase chain reaction (qRT-PCR).
H1299 and A549 cells were transfected with negative control (NC) or miR-1247 mimics (HmiR0610, Funeng, Guangzhou, People’s Republic of China) using Lip2000 (Thermo Fisher Scientific) for 48 h. A group of cells (5-Aza group) were treated with 5 μmol/L of 5-Aza for 48 h. All the cells were examined for the ability to invade and metastasize by Western blot and methylation-specific PCR (MSP).

Zhang J., Fu J., Pan Y., Zhang X, & Shen L. (2016). Silencing of miR-1247 by DNA methylation promoted non-small-cell lung cancer cell invasion and migration by effects of STMN1. OncoTargets and therapy, 9, 7297-7307.

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Demethylation of GBGT1 Gene

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Cells carrying a hypermethylated GBGT1 gene were treated with 5-Aza (Sigma-Aldrich Pty. Ltd; stock solution prepared in 50% (v/v) acetic acid) in order to restore or increase GBGT1 expression. An amount of 10⁵ cells were seeded in 6-well plates (NUNC, Thermo Fisher Scientific Pty. Ltd., Scoresby, Australia) and treated with 5-Aza (2.5 μM final concentration) on the next day. The culture medium was removed every 24 h and replaced by fresh culture medium containing 2.5 μM 5-Aza. Samples were harvested after 24 h, 48 h and 72 h of treatment. Genomic DNA and total RNA were extracted as described above. Mock controls contained 50% (v/v) acetic acid at concentrations identical to 5-Aza.

Jacob F., Hitchins M.P., Fedier A., Brennan K., Nixdorf S., Hacker N.F., Ward R, & Heinzelmann-Schwarz V.A. (2014). Expression of GBGT1 is epigenetically regulated by DNA methylation in ovarian cancer cells. BMC Molecular Biology, 15, 24.

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Cytotoxic Effects of 5-AZA on Colorectal Cancer

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Four CRC cell lines and normal NCM460 cells were seeded in 96-well plates at a density of 5 × 10³ to a final volume of 200 μl and treated with various concentrations (0.1, 0.5, 1, 2, 5, 10, 20 μM) of 5-AZA (Sigma Aldrich cat#A3656) diluted in phosphate-buffered saline (PBS) (Gibco cat#10010023) for 48 h. A negative control (without 5-AZA) and a blank control (without cells) were included in each plate. At the end of treatment course, Cell Counting Kit-8 assay (Dojindo cat#CK04) was used to exam the cell proliferation status. The optical density (OD) was determined at 490 nm using an ELISA reader (BIORAD). Cell viability = (OD of treated-OD of brank)/(OD of the negative control-OD of the blank) × 100%.
As described above, four CRC cell lines and normal NCM460 cells were seeded in six-well plate and exposed to 5-AZA at a concentration of 5 μM for 48 h. The control group was treated in parallel with PBS. DNA, RNA and protein were harvested for downstream analysis. Each sample was tested in triplicate.

Zhou D., Tang W., Su G., Cai M., An H.X, & Zhang Y. (2017). PCDH18 is frequently inactivated by promoter methylation in colorectal cancer. Scientific Reports, 7, 2819.

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Ginsenoside Effects on MCF-7 Breast Cancer Cells

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The breast cancer cell line MCF-7 was purchased from the American Type Culture Collection (ATCC; Manassas, VA, USA). The cells were cultured in RPMI 1640 medium (Gibco BRL, Carlsbad, CA, USA) supplemented with 10% FBS (Capricorn, Ebsdorfergrund, Germany) and 1% penicillin-streptomycin. The ginsenoside Rg3 and Rh2 (LKT Labs, St. Paul, Minnesota, USA), Korean Red Ginseng (KRG) extract (kindly donated by Korea Ginseng Co., Daejon, Korea), and 5-Aza-2′-deoxycytidine (5-Aza; Sigma-Aldrich, St. Louis, MO, USA) were used to treat the cells in a 60-mm culture dish at a confluence of 50-70%. The contents of various ginsenosides in KRG was determined as described previously [18] (link). Rg3 and Rh2 stocks were prepared with 20 mM in 100% ethanol and used in final concentrations of 20 and 50 μM. KRG extract was dissolved in distilled water and treated to cells at final concentrations of ∼200 μg/ml. 5-Aza stock was prepared with 10 mM in 100% DMSO (Sigma-Aldrich) and treated to cells in final concentrations of 2.5, 5, and 10 μM. Twenty-four hours after treatment by Rg3, Rh2, KRG extract, or 5-Aza, the cells were harvested, washed with PBS, and trypsinized using 0.05% trypsin-EDTA (Gibco BRL).

Ham J., Jeong D., Park S., Kim H.W., Kim H, & Kim S.J. (2019). Ginsenoside Rg3 and Korean Red Ginseng extract epigenetically regulate the tumor-related long noncoding RNAs RFX3-AS1 and STXBP5-AS1. Journal of Ginseng Research, 43(4), 625-634.

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