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Visualization of Notch Signaling Oscillation in Cells and Tissues

  • Hiromi Shimojo
  • Yukiko Harima
  • Ryoichiro KageyamaEmail author
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1187)

Abstract

The Notch signaling effectors Hes1 and Hes7 exhibit oscillatory expression with a period of about 2–3 h during embryogenesis. Hes1 oscillation is important for proliferation and differentiation of neural stem cells, whereas Hes7 oscillation regulates periodic formation of somites. Continuous expression of Hes1 and Hes7 inhibits these developmental processes. Thus, expression dynamics are very important for gene functions, but it is difficult to distinguish between oscillatory and persistent expression by conventional methods such as in situ hybridization and immunostaining. Here, we describe time-lapse imaging methods using destabilized luciferase reporters and a highly sensitive cooled charge-coupled device camera, which can monitor dynamic gene expression. Furthermore, the expression of two genes can be examined simultaneously by a dual reporter system using two-color luciferase reporters. Time-lapse imaging analyses reveal how dynamically gene expression changes in many biological events.

Key words

Hes1 Hes7 Oscillatory expression Luciferase Bioluminescence CCD Neural stem cell Presomitic mesoderm Segmentation clock 

Notes

Acknowledgements

This work was supported by Core Research for Evolutional Science and Technology and Grant-in-Aid for Scientific Research on Innovative Areas (MEXT 22123002).

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Copyright information

© Springer Science+Business Media New York 2014

Authors and Affiliations

  • Hiromi Shimojo
    • 1
  • Yukiko Harima
    • 2
  • Ryoichiro Kageyama
    • 3
    Email author
  1. 1.Institute for Virus Research and World Premier International Research Initiative–Institute for Integrated Cell-Material Sciences (WPI-iCeMS)Kyoto UniversityKyotoJapan
  2. 2.Institute for Virus ResearchKyoto University, and Core Research for Evolutional Science and Technology (CREST), Japan Science and Technology AgencyKyotoJapan
  3. 3.Institute for Virus Research and World Premier International Research Initiative–Institute for Integrated Cell-Material Sciences (WPI-iCeMS)Kyoto University, and Core Research for Evolutional Science and Technology (CREST), Japan Science and Technology AgencyKyotoJapan

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