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Wnt Signaling pp 141–159Cite as

The Generation of Organoids for Studying Wnt Signaling

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Part of the book series: Methods in Molecular Biology ((MIMB,volume 1481))

Abstract

We established an in vitro culture model in which intestinal epithelial stem cells can grow into three-dimensional, ever-expanding epithelial organoids that retain their original organ identity and genetic stability. Moreover, organoids can easily be genetically modified using different genome modification strategies, including viral delivery of transgenes and CRISPR/Cas9 technology. These combined characteristics make them a useful in vitro model system to study many biological processes including the contribution of cellular signaling pathways to tissue homeostasis and disease. Here we describe our current laboratory protocols to establish human intestinal organoids and how to genetically modify both mouse and human intestinal organoids to study cellular signaling pathways, specifically Wnt signaling. Moreover, we provide a detailed protocol for lentiviral transduction and CRISPR/Cas9-mediated genome modification of organoid cultures.

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Acknowledgements

We thank Johan H. van Es for critical reading of the manuscript. We are grateful for support from the following: The Netherlands Organisation for Scientific Research (NWO-ZonMw) VENI grant to J.D. (91614138); FEBS long term fellowship to B.A.

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Authors and Affiliations

  1. Hubrecht Institute-KNAW and University Medical Center Utrecht, Uppsalalaan 8, 3584 CT, Utrecht, The Netherlands

    Jarno Drost, Benedetta Artegiani & Hans Clevers

Authors
  1. Jarno Drost
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  2. Benedetta Artegiani
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  3. Hans Clevers
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Corresponding author

Correspondence to Hans Clevers .

Editor information

Editors and Affiliations

  1. University of Texas Southwestern Medical Center, Dallas, Texas, USA

    Quinn Barrett

  2. Department of Cell Biology, University of Texas Southwestern Medical Center, Dallas, Texas, USA

    Lawrence Lum

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Drost, J., Artegiani, B., Clevers, H. (2016). The Generation of Organoids for Studying Wnt Signaling. In: Barrett, Q., Lum, L. (eds) Wnt Signaling. Methods in Molecular Biology, vol 1481. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-6393-5_15

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  • DOI: https://doi.org/10.1007/978-1-4939-6393-5_15

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  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-6391-1

  • Online ISBN: 978-1-4939-6393-5

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