Gfi 1

Manufactured by Santa Cruz Biotechnology

Sourced in Canada, United States

Gfi-1 is a DNA-binding transcriptional repressor that plays a role in the regulation of gene expression. It is an important regulator of hematopoietic and neural development.

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Lab products found in correlation

4 protocols using gfi 1

Western Blot Analysis of Nuclear Proteins

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Cell nuclear lysates were isolated using the NE-PER extraction kit (Thermo Scientific, Rockford, IL). Equal amounts of protein lysate (15 μg) were resolved by SDS–PAGE and transferred onto PVDF membrane (Millipore, Billerica, MA). Blots were blocked and incubated with primary and secondary antibody as described previously [18] (link). Primary antibodies were specific against C/EBPα, RARα, VDR, EGR1, PU.1, Oct4 (Cell Signaling, Danvers, MA), AhR, Gfi-1 (Santa Cruz Biotechnology, Santa Cruz, CA) and IRF-1 (BD Biosciences). Anti-Histone 3 or anti-TATA-binding protein (Cell Signaling) were used to ensure even loading.

Jensen H.A., Yourish H.B., Bunaciu R.P., Varner J.D, & Yen A. (2015). Induced myelomonocytic differentiation in leukemia cells is accompanied by noncanonical transcription factor expression. FEBS Open Bio, 5, 789-800.

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Antibody Panel for Western Blot Analysis

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Antibodies used for Western blot analyses were as follows: actin (Sigma-Aldrich, no. A4700, clone AC-40), GFI1 (Santa Cruz Biotechnology, sc-376949, clone B9), GSE1 (Proteintech, no. 24947-I-AP), H3 (Abcam, no. 1791), H3K4me1 (Abcam, no. 8895), H3K4me2 (Abcam, no. 32356), H3K4me3 (Active Motif, no. 39159), HDAC1 (Abcam, no. 7028), HMG20B (Proteintech, no. 14582-1-AP), laminin B (Abcam, no. 16048), LSD1 (Cell Signaling Technology, no. 2139 and Abcam, no. 17721), RAR (Santa Cruz Biotechnology, sc-551), tubulin (Santa Cruz Biotechnology, sc-32356), vinculin (Sigma-Aldrich, no. V9131, clone hVIN-1), and ZMYM3 (Abcam, no. 106626).

Ravasio R., Ceccacci E., Nicosia L., Hosseini A., Rossi P.L., Barozzi I., Fornasari L., Zuffo R.D., Valente S., Fioravanti R., Mercurio C., Varasi M., Mattevi A., Mai A., Pavesi G., Bonaldi T, & Minucci S. (2020). Targeting the scaffolding role of LSD1 (KDM1A) poises acute myeloid leukemia cells for retinoic acid–induced differentiation. Science Advances, 6(15), eaax2746.

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ChIP Assay for miR-22 Regulation

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ChIP assay was performed, as described previously14 (link)17 (link), with SABiosciences Corporation’s ChampionChiP One-Day kit (Qiagen, Frederick, MD) following the manufacturer’s protocol, with some modifications. Briefly, pellets of 5 × 10⁶ cells were treated with fresh fixing buffer (1% formaldehyde) for 10 min at 37 °C to crosslink DNA and proteins. The reaction was terminated by the addition of stop buffer and incubated at room temperature for 5 min. After cell lysis, the cross-linked chromatin was sonicated to an average size of ∼500 bp and was immunoprecipitated with antibodies against TET1, GFI1 (Santa Cruz Biotechnology Inc., Santa Cruz, CA), the N′-terminal portion of MLL (MLL-N), the C′-terminal of MLL (MLL-C), H3K27Me3, H3K4Me3, RNA polymerase II, EZH2, SIN3A or IgG (Abcam, Cambridge, MA). Purified ChIP DNA was amplified by real-time qPCR using specific primers targeting the CpG-enriched upstream region of human miR-22: forward: 5′- GTTGTTGGAGTCGTGAGTG -3′; reverse: 5′- CGCTCCACCTTTCCTTAAA -3′; or mouse miR-22: forward: 5′- TGAATGGGCGGGAGTAA -3′; reverse: 5′- CCACGAGCTGCGAATGAA -3′.

Jiang X., Hu C., Arnovitz S., Bugno J., Yu M., Zuo Z., Chen P., Huang H., Ulrich B., Gurbuxani S., Weng H., Strong J., Wang Y., Li Y., Salat J., Li S., Elkahloun A.G., Yang Y., Neilly M.B., Larson R.A., Le Beau M.M., Herold T., Bohlander S.K., Liu P.P., Zhang J., Li Z., He C., Jin J., Hong S, & Chen J. (2016). miR-22 has a potent anti-tumour role with therapeutic potential in acute myeloid leukaemia. Nature Communications, 7, 11452.

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Apoptosis Signaling Pathway Inhibitors

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UA (C₃₀H₄₈O₃, MW = 456.68) was obtained from Sigma Chemical Co., St. Louis, USA. SH-4-54 was from Selleck Chemicals, Houston, USA. Imatinib (Imatinib Mesylate, Gleevec, STI571) was from Novartis Pharmaceuticals Basel, Switzerland. All reagents were prepared and used as recommended by their suppliers. Antibodies against PARP, Bcl-xL, Mcl-1, Bad, p-Stat5 at Tyr⁶⁹⁴, p-Akt at Ser⁴⁷³, p-Akt at Thy³⁰⁸, Akt, Akt1, Akt2, Procaspase9, Caspase3, c-Abl, and p-c-Abl at Tyr²⁴⁵ were from Cell Signaling Technology, Beverly, USA. The Antibody against Stat5 was from Zymed Laboratories Inc., South San Francisco, USA. Antibodies against p-Bad at Ser¹³⁶, Gfi-1, and β-actin and HRP-conjugated secondary antibodies were from Santa Cruz Biotechnology Inc., Santa Cruz, USA.

Lin Z., Jiang J, & Liu X.S. (2016). Ursolic acid-mediated apoptosis of K562 cells involves Stat5/Akt pathway inhibition through the induction of Gfi-1. Scientific Reports, 6, 33358.

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