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Luciferase reporter assay.

Frank R Wettey1, Antony P Jackson

  • 1Department of Biochemistry, University of Cambridge, Tennis Court Road, Cambridge CB2 1QW, UK.

Sub-Cellular Biochemistry
|July 13, 2007
PubMed
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This study presents a simple, sensitive luciferase reporter assay to measure Tet-system responsiveness in DT40 cells. The assay is quantitative, reproducible, and suitable for high-throughput screening using transient or stable transfections.

Area of Science:

  • Biochemistry
  • Molecular Biology
  • Cell Biology

Background:

  • The Tet-system is a widely used inducible gene expression system.
  • Assessing Tet-system responsiveness is crucial for experimental design and data interpretation.
  • Existing methods may lack sensitivity, reproducibility, or ease of use.

Purpose of the Study:

  • To develop and validate a simple, highly sensitive luciferase reporter assay.
  • To establish a quantitative and reproducible method for determining Tet-system responsiveness.
  • To demonstrate the assay's utility in DT40 cell screens with various transfection types.

Main Methods:

  • Utilized firefly luciferase as a reporter gene.
  • Measured photon emission from luciferin oxidation catalyzed by luciferase.

Related Experiment Videos

  • Quantified light output as relative light units (RLU) using a luminometer.

    • Main Results:

    • The luciferase reporter assay demonstrated high sensitivity and responsiveness to the Tet-system.
    • The assay proved to be quantitative and reproducible across experiments.
    • Successfully applied the assay in DT40 cell screens involving both transient and stable transfections.

    Conclusions:

    • The developed luciferase reporter assay offers a robust and user-friendly method for assessing Tet-system responsiveness.
    • This assay is well-suited for high-throughput screening applications in DT40 cells.
    • Provides a reliable tool for researchers working with inducible gene expression systems.