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Gastruloids: Embryonic Organoids from Mouse Embryonic Stem Cells to Study Patterning and Development in Early Mammalian Embryos

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Programmed Morphogenesis

Part of the book series: Methods in Molecular Biology ((MIMB,volume 2258))

Abstract

Gastruloids are embryonic organoids made from small, defined numbers of mouse embryonic stem cells (mESCs) aggregated in suspension culture, which over time form 3D structures that mimic many of the features of early mammalian development. Unlike embryoid bodies that are usually disorganized when grown over several days, gastruloids display distinct, well-organized gene expression domains demarcating the emergence of the three body axes, anteroposterior axial elongation, and implementation of collinear Hox transcriptional patterns over 5–7 days of culture. As such gastruloids represent a useful experimental system that is complementary to in vivo approaches in studying early developmental patterning mechanisms regulating the acquisition of cell fates. In this protocol, we describe the most recent method for generating gastruloids with high reproducibility, and provide a comprehensive list of possible challenges as well as steps for protocol optimization.

The former name of the author “Peter Baillie-Benson” was “Peter Baillie-Johnson”.

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Acknowledgments

The authors wish to thank members of Alfonso Martinez Arias’ lab, University of Cambridge, UK, for discussions. Additionally, for kindly sharing their cell lines, we are indebted to Heiko Lickert (FoxA2H2BYFP), Gordon Keller (Bra+/GFP), Austin Smith (E14Tg2A, 46C Sox1GFP), Palle Serup (AR8mCherry), Jérôme Collignon (Nodal+/YFP), Anna-Katerina Hadjantonakis (GATA6H2BVenus), and David Suter (Sox1eGFP; BramCherry). We thank K. Hötte-Lohmeier, F. Pampaloni, and E.H.K. Stelzer from Goethe Universität Frankfurt am Main, Germany, for providing collectors and multi-collectors. KA, KA, and VT are funded by the European Molecular Biology Laboratory (EMBL) Barcelona; P B-B is funded as part of a Wellcome Strategic Award to Professor Jennifer Nichols and a European Research Council advanced grant to Professor Alfonso Martinez Arias (834580). The Cambridge Stem Cell Institute is supported by core funding from Wellcome and the Medical Research Council. DAT is funded by an NC3Rs David Sainsbury Research Fellowship (NC/P001467/1), a Wellcome Trust Institutional Strategic Support Fund (ISSF), and by an award from the University of Liverpool Technology Directorate Voucher Scheme. We acknowledge the Liverpool Centre for Cell Imaging (CCI) for provision of imaging equipment and technical assistance. Kerim Anlas and Peter Baillie-Benson contributed equally to this work.

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Author notes

  1. Kerim Anlas and Peter Baillie-Benson authors contributed equally to this work.

Authors and Affiliations

  1. European Molecular Biology Laboratory (EMBL), Barcelona, Barcelona, Spain

    Kerim Anlas, Krisztina Arató & Vikas Trivedi

  2. EMBL Heidelberg, Developmental Biology Unit, Heidelberg, Germany

    Kerim Anlas

  3. Formerly Wellcome—MRC Cambridge Stem Cell Institute, Department of Genetics, University of Cambridge, Cambridge, UK

    Peter Baillie-Benson

  4. Institute of Life Course and Medical Sciences, University of Liverpool, Liverpool, UK

    David A. Turner

Authors
  1. Kerim Anlas
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  2. Peter Baillie-Benson
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  3. Krisztina Arató
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  4. David A. Turner
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  5. Vikas Trivedi
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Corresponding authors

Correspondence to David A. Turner or Vikas Trivedi .

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Editors and Affiliations

  1. Department of Pathology, Division of Experimental Pathology, School of Medicine, University of Pittsburgh, Pittsburgh Liver Research Center, University of Pittsburgh, Department of Bioengineering Swanson School of Engineering, University of Pittsburgh, McGowan Institute for Regenerative Medicine, University of Pittsburgh, Pittsburgh, PA, USA

    Mo R. Ebrahimkhani

  2. Department of Bioengineering Swanson School of Engineering, Pittsburgh Liver Research Center, Department of Pathology, Division of Experimental Pathology, School of Medicine, University of Pittsburgh, Pittsburgh, PA, USA

    Joshua Hislop

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Anlas, K., Baillie-Benson, P., Arató, K., Turner, D.A., Trivedi, V. (2021). Gastruloids: Embryonic Organoids from Mouse Embryonic Stem Cells to Study Patterning and Development in Early Mammalian Embryos. In: Ebrahimkhani, M.R., Hislop, J. (eds) Programmed Morphogenesis. Methods in Molecular Biology, vol 2258. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-1174-6_10

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  • DOI: https://doi.org/10.1007/978-1-0716-1174-6_10

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  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-0716-1173-9

  • Online ISBN: 978-1-0716-1174-6

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