Dtag 13

Manufactured by Merck Group

DTAG-13 is a laboratory equipment product manufactured by Merck Group. It is designed for precise measurement and analysis applications within controlled laboratory settings. The core function of DTAG-13 is to provide accurate and reliable data collection capabilities for scientific research and testing purposes.

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Lab products found in correlation

7 protocols using dtag 13

Protein Degradation Assay in Mammalian Cells

Cited 1 time

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Cells were seeded onto 8-chamber coverglass (Nunc^™ Lab-Tek^™ II Chambered Coverglass, 155409) coated with 5 μg/ml laminin (BioLamina LN511), in 2i media and appropriate drugs. For all experiments, cells were grown overnight in the chambers. Protein degradation was induced by treating cells with a final concentration of 400 μM dTAG-13 (Sigma SML2601) for 2 hrs or 6 hrs (2 hrs: Sox2, Taf1, Tbp/Trf2, Gtf2f1, CDK7; 6 hrs: Rad21 and Med19). 0.04% v/v DMSO was used as control. Before imaging, cells were labeled with the relevant Halo-dye (Promega, Janelia Fluor^® HaloTag^® Ligands) or SNAP-dye (SNAP-Cell^® 647-SiR, NEB S9102S) with media containing 0.3 μM dye for 10 min, at 37°C, followed by three times rinsing with new media. Rpb1 protein degradation was induced by treating cells with 40 nM Halo-PROTAC-E(34 (link)) for 2 hrs and 5 nM Halo-dye for 1 hr. For THZ1 and Triptolide treatment, cells were treated by 100 nM for 1hr. For combined dTAG and inhibitor treatment, cells were treated by dTAG-13 for 6 hrs and 100 nM inhibitor for 1hr.

Cheng L., De C., Li J, & Pertsinidis A. (2023). Mechanisms of transcription control by distal enhancers from high-resolution single-gene imaging. bioRxiv.

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Chemical Agents for Cell Biology Research

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Doxycycline was used at 1 μg/ml (Sigma), Indole-3-acetic acid (IAA) used at 500 μM (Sigma, I5148), CDK1 inhibitor RO-3306 used at 9 μM (Sigma-Aldrich, SML0569), NMS-P715 0.8 μM (Millipore Sigma, 475949), ICRF-193 used at 100 nM (Enzo life sciences, BML-GR332–0001) S-Trityl-L-cysteine (STLC) used at 5 μM (Enzo Life Sciences, ALX-105–011-M500), dTAG-13 used at 500 nM (Sigma-Aldrich, SML2601), Okadaic acid used at 500 nM (Fisher Scientific, 11–362-5U), Geneticin^® Selective Antibiotic (G418 Sulfate) used at 300 μg/ml (Gibco, 10131035).

Broberger C., Johansen J., Johansson C., Schalling M, & Hökfelt T. (1998). The neuropeptide Y/agouti gene-related protein (AGRP) brain circuitry in normal, anorectic, and monosodium glutamate-treated mice. Proceedings of the National Academy of Sciences of the United States of America, 95(25), 15043-15048.

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dTAG-13 Protein Degradation Assay

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dTAG-13 (Bio-Techne) was reconstituted in DMSO (Sigma) at 5mM. dTAG-13 was diluted in maintenance medium to 500nM and added to cells with medium changes for the specified amounts of time.

Abuhashem A., Lee A.S., Joyner A.L, & Hadjantonakis A.K. (2022). Rapid and efficient degradation of endogenous proteins in vivo identifies stage specific roles of RNA Pol II pausing in mammalian development. Developmental cell, 57(8), 1068-1080.e6.

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Targeted degradation of NELFB in mECSs

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Cells were cultured in a humidified 37°C incubator with 5% CO2. K562 (ATCC, CCL-243) and Jurkat (ATCC, TIB-152) cells were grown in RPMI-1640 medium supplemented with 10% fetal bovine serum and 1X penicillin streptomycin antibiotic.
mECSs (Mouse embryonic stem cell line E14, ATCC, CRL-1821) harboring a homozygous endogenous NELFB-FKBP12^F36V fusion protein^{61 (link),84 (link)} were cultured on 0.1% gelatin (Millipore) in PBS+/+ coated tissue-culture grade plates. For routine culture, cells were grown in Serum/LIF conditions: DMEM (Gibco), supplemented with 2 mM L-glutamine (Gibco), 1x MEM non-essential amino acids (Gibco), 1 mM sodium pyruvate (Gibco), 100 U/ml penicillin and 100 U/ml streptomycin (Gibco), 0.1 mM 2-mercaptoethanol (Gibco), 15% fetal bovine serum (Gibco), and 1000 U/ml of recombinant leukemia inhibitory factor (LIF).
To induce NELFB degradation, dTAG-13 (Bio-Techne) was reconstituted in DMSO (Sigma) at 5 mM. dTAG-13 was diluted in maintenance medium to 500 nM and added to cells with medium changes for the specified amounts of time. For dTAG washes, the cells were washed 4 times, twice with PBS +/+ and twice with maintenance medium following the treatment time to ensure complete removal of the dTAG ligand. At the end of each dTAG-13 treatment time point, cells were detached using Trypsin-EDTA (0.05%) (Gibco) and counted before crosslinking for Micro-C.

Ward D.M., Pevsner J., Scullion M.A., Vaughn M, & Kaplan J. (2000). Syntaxin 7 and VAMP-7 are Soluble N-Ethylmaleimide–sensitive Factor Attachment Protein Receptors Required for Late Endosome–Lysosome and Homotypic Lysosome Fusion in Alveolar Macrophages. Molecular Biology of the Cell, 11(7), 2327-2333.

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Immunomodulatory Drugs for Cell Assays

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IMiD compounds (Thalidomide, Lenalidomide, Pomalidomide and Iberdomide) were obtained from Caymen Chemicals. BTX161 compound was synthesised in-house and is available for request from the MRC-PPU reagents website (http://mrcppureagents.dundee.ac.uk). IMiD compounds were added to cell culture media at indicated concentrations (between 0.1 and 10 μM) for indicated duration. dTAG-13 (Sigma-Aldrich), a CRBN-binding PROTAC, was used at 1 μM for 24 h. MLN4924 (Sigma-Aldrich), an inhibitor of NEDD8-activating E1 enzyme, was used at 1 μM for 24 h. Bortezomib (Sigma-Aldrich), a proteasome inhibitor, was used at 5 μM for 24 h.

Dunbar K., Macartney T.J, & Sapkota G.P. (2020). IMiDs induce FAM83F degradation via an interaction with CK1α to attenuate Wnt signalling. Life Science Alliance, 4(2), e202000804.

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Modulating Protein Degradation with dTAG-13

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dTAG-13 (Bio-Techne) was reconstituted in DMSO (Sigma) at 5 mM. dTAG-13 was diluted in maintenance medium to 500 nM and added to cells with medium changes for the specified amounts of time.

Abuhashem A., Chivu A.G., Zhao Y., Rice E.J., Siepel A., Danko C.G, & Hadjantonakis A.K. (2022). RNA Pol II pausing facilitates phased pluripotency transitions by buffering transcription. Genes & Development, 36(13-14), 770-789.

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dTAG-13 Compound Treatment Protocol

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Cells dTAG treatment dTAG-13 (Bio-Techne) was reconstituted in DMSO (Sigma) at 5mM. dTAG-13 was diluted in maintenance medium to 500nM and added to cells with medium changes for the specified amounts of time.

Abuhashem A., & Hadjantonakis A. (2021). Rapid and efficient adaptation of the dTAG system in mammalian development reveals stage specific requirements of NELF.

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