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Analysis of Signaling Pathways in Zebrafish Development by Microinjection

  • William H. Kinsey
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 518)

Abstract

The zebrafish oocyte differs substantially from the zygote and cleavage-stage embryo with regard to the ease with which it can be microinjected with proteins or reagents that modify subsequent development. The objective of this chapter is to describe methods developed in this and other laboratories for microinjection and calcium imaging in the unfertilized zebrafish egg. Methods of immobilizing the oocyte include a holding chamber and a holding pipette. The holding chamber allows imaging of three or four oocytes simultaneously, while the holding pipette facilitates imaging of localized regions in the oocyte. Injection of calcium green dextran via holding chambers allowed detection of global changes in Ca2+ release following fertilization and development through early blastula stages. Injection and imaging with the holding pipette method allowed discrimination of calcium changes in the egg cortex from that in the central regions of the cell. The results demonstrate the highly localized nature of calcium signaling in the zebrafish zygote and the implications of this signaling for embryonic development.

Key words

Zebrafish egg oocyte zygote fertilization Fyn Yes Src 

Notes

Acknowledgments

Supported by NICHD-HD14846.

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Copyright information

© Humana Press, a part of Springer Science+Business Media, LLC 2009

Authors and Affiliations

  • William H. Kinsey
    • 1
  1. 1.Department of Anatomy and Cell BiologyUniversity of Kansas Medical CenterKansas CityUSA

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