Quantifying NOTCH Transcriptional Activity

NOTCH transcriptional activity was analyzed using luciferase assays, as previously described [18 (link),19 (link),22 (link),84 (link)]. We also treated C3H10T1/2 cells with the γ-secretase inhibitor DAPT (N-[N-(3,5-Difluorophenacetyl)-L-alanyl]-S-phenylglycine t-butyl ester) (10 μM), used as a NOTCH-signaling-inhibition control. Luciferase assays were measured using the Orion II microplate luminometer (Berthold), and samples were processed with the Dual-Luciferase Reporter Assay System (Promega), following the supplier’s recommendations. Assays were repeated at least three times.

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Rodríguez-Cano M.M., González-Gómez M.J., Sánchez-Solana B., Monsalve E.M., Díaz-Guerra M.J., Laborda J., Nueda M.L, & Baladrón V. (2020). NOTCH Receptors and DLK Proteins Enhance Brown Adipogenesis in Mesenchymal C3H10T1/2 Cells. Cells, 9(9), 2032.

Publication 2020

Orion II microplate luminometer Dual-Luciferase Reporter Assay System

Assays Cells Dapt Ester Inhibition Luciferase Promega Secretase Transcriptional

Corresponding Organization : University of Castilla-La Mancha

Other organizations : National Cancer Institute, National Institutes of Health, Center for Cancer Research

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Variable analysis

independent variables

Treatment with the γ-secretase inhibitor DAPT (N-[N-(3,5-Difluorophenacetyl)-L-alanyl]-S-phenylglycine t-butyl ester) (10 μM)

dependent variables

NOTCH transcriptional activity analyzed using luciferase assays

control variables

Luciferase assays were measured using the Orion II microplate luminometer (Berthold)
Samples were processed with the Dual-Luciferase Reporter Assay System (Promega)

positive control

NOTCH-signaling-inhibition control

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