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Generation of Ectopic Morphogen Gradients in the Zebrafish Blastula

  • Maraysa de Olivera-Melo
  • Peng-Fei Xu
  • Nathalie Houssin
  • Bernard Thisse
  • Christine Thisse
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1863)

Abstract

In the zebrafish embryo, cells of the early blastula animal pole are all equivalent and are fully pluripotent until the midblastula transition that occurs at the tenth cell cycle (512 to 1K cells). This naive territory of the embryo is therefore perfectly suited to assay for morphogen activity. Here we describe different methods to generate ectopic morphogen gradients, either in vivo at the animal pole of the embryo, or in vitro in animal pole explants or in aggregates of animal pole blastomeres (also named embryoid bodies). These methods include injection of mRNA coding for growth factor(s) into animal pole blastomere(s), transplantation of growth factor(s) secreting cells, implantation of beads coated with purified protein(s), and various combinations of these different approaches. Our comparative study reveals that all these methods allow to generate morphogen gradient(s) that are able to induce, both in vivo and in vitro, the formation of a well-patterned embryonic axis.

Key words

Morphogen Gradient Zebrafish Blastula Injection Cell transplantation Bead implantation Embryoid bodies 

Notes

Acknowledgments

This work was supported by funds from University of Virginia (B.T., C.T.), March of Dimes (1-FY15-298 to B.T.), Jefferson Trust (FAAJ3199 to C.T.), Conselho Nacional de Desenvolvimento Científico e Tecnológico, Brazil (grant number 200535/2014-5 to M. de O-M.).

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Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2018

Authors and Affiliations

  • Maraysa de Olivera-Melo
    • 1
  • Peng-Fei Xu
    • 1
  • Nathalie Houssin
    • 1
  • Bernard Thisse
    • 1
  • Christine Thisse
    • 1
  1. 1.Department of Cell BiologyUniversity of Virginia School of MedicineCharlottesvilleUSA

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