Reporter Assays

  • M. Fernanda Ceriani
Part of the Methods in Molecular Biology™ book series (MIMB, volume 362)


Transcriptional feedback loops are at the core of the molecular clockworks. As single clock genes were cloned it was compelling to develop an assay that allowed simple and direct functional testing of putative activators or repressors of transcription. This chapter includes a general description and guidelines to carry out transcriptional assays in transiently transfected Schneider’s cells.

Key Words

Transcriptional assays firefly luciferase Renilla luciferase S2 cells 


  1. 1.
    Darlington, T. K., Wager-Smith, K., Ceriani, M. F., et al. (1998) Closing the circadian loop: CLOCK-induced transcription of its own inhibitors per and tim. Science 280, 1599–1603.CrossRefPubMedGoogle Scholar
  2. 2.
    Froy, O., Chang, D.C., and Reppert, S.M. (2002) Redox potential: differential roles in dCRY and mCRY1 functions. Curr. Biol. 12, 147–152.CrossRefPubMedGoogle Scholar
  3. 3.
    Chang, D. C., McWatters, H. G., Williams, J. A., Gotter, A. L., Levine, J. D., and Reppert, S. M. (2003) Constructing a feedback loop with circadian clock molecules from the silkmoth, Antheraea pernyi. J. Biol. Chem. 278, 38,149–38,158.CrossRefPubMedGoogle Scholar
  4. 4.
    Ceriani, M. F., Darlington, T. K., Staknis, D., et al. (1999). Light-dependent sequestration of TIMELESS by CRYPTOCHROME. Science 285, 553–556.CrossRefPubMedGoogle Scholar
  5. 5.
    Ko, H. W., Jiang, J., and Edery, I. (2002) Role for Slimb in the degradation of Drosophila Period protein phosphorylated by Doubletime. Nature 420, 673–678.CrossRefPubMedGoogle Scholar
  6. 6.
    Heim, R., Prasher, D. C., and Tsien, R. Y. (1994) Wavelength mutations and posttranslational autoxidation of green fluorescent protein. Proc. Natl. Acad. Sci. USA 91, 12,501–12,504.CrossRefPubMedGoogle Scholar
  7. 7.
    von Ohlen, T., Lessing, D., Nusse, R., and Hooper, J. E. (1997) Hedgehog signaling regulates transcription through cubitus interruptus, a sequence-specific DNA binding protein. Proc. Natl. Acad. Sci. USA 94, 2404–2409.CrossRefGoogle Scholar
  8. 8.
    Brandes, C., Plautz, J. D., Stanewsky, R., et al. (1996) Novel features of Drosophila period transcription revealed by real-time luciferase reporting. Neuron 16, 687–692.CrossRefPubMedGoogle Scholar
  9. 9.
    Plautz, J. D., Straume, M., Stanewsky, R., et al. (1997) Quantitative analysis of Drosophila period gene transcription in living animals. J. Biol. Rhythms 12, 204–217.CrossRefPubMedGoogle Scholar
  10. 10.
    Stanewsky, R., Jamison, C. F., Plautz, J. D., Kay, S. A., and Hall, J. C. (1997) Multiple circadian-regulated elements contribute to cycling period gene expression in Drosophila. EMBO J. 16, 5006–5018.CrossRefPubMedGoogle Scholar
  11. 11.
    Plautz, J. D., Kaneko, M., Hall, J.C., and Kay, S. A. (1997) Independent photoreceptive circadian clocks throughout Drosophila. Science 278, 1632–1635.CrossRefPubMedGoogle Scholar
  12. 12.
    Hao, H., Allen, D. L. and Hardin, P. E. (1997) A circadian enhancer mediates PER-dependent mRNA cycling in Drosophila melanogaster. Mol. Cell Biol. 17, 3687–3693.PubMedGoogle Scholar
  13. 13.
    Froy, O., Chang, D.C., and Reppert, S. M. (2002) Redox potential: differential roles in dCRY and mCRY1 functions. Curr. Biol. 12, 147–152.CrossRefPubMedGoogle Scholar
  14. 14.
    Sonnenfeld, M., Ward, M., Nystrom, G., Mosher, J., Stahl, S., and Crews, S. (1997) The Drosophila tango gene encodes a bHLH-PAS protein that is orthologous to mammalian Arnt and controls CNS midline and tracheal development. Development 124, 4571–4582.PubMedGoogle Scholar
  15. 15.
    Nawathean, P., and Rosbash, M. (2004) The doubletime and CKII kinases collaborate to potentiate Drosophila PER transcriptional repressor activity. Mol. Cell 13, 213–223.CrossRefPubMedGoogle Scholar
  16. 16.
    Weber, F., and Kay, S. A. (2003) A PERIOD inhibitor buffer introduces a delay mechanism for CLK/CYC-activated transcription. FEBS Lett. 555, 341–345.CrossRefPubMedGoogle Scholar

Copyright information

© Humana Press Inc. 2007

Authors and Affiliations

  • M. Fernanda Ceriani
    • 1
  1. 1.Department Behavioral GeneticsFundación Instituto LeloirBuenos AiresArgentina

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