A functional genetic toolbox for human tissue-derived organoids.
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Authors
Sun, Dawei
Perrone, Francesca
Sokleva, Vanesa
Lim, Kyungtae
Rezakhani, Saba
Lutolf, Matthias
Zilbauer, Matthias
Publication Date
2021-10-06Journal Title
eLife
ISSN
2050-084X
Volume
10
Language
eng
Type
Article
This Version
VoR
Metadata
Show full item recordCitation
Sun, D., Evans, L., Perrone, F., Sokleva, V., Lim, K., Rezakhani, S., Lutolf, M., et al. (2021). A functional genetic toolbox for human tissue-derived organoids.. eLife, 10 https://doi.org/10.7554/elife.67886
Description
Funder: Alzheimers Research UK Stem Cell Research Centre
Abstract
Human organoid systems recapitulate key features of organs offering platforms for modelling developmental biology and disease. Tissue-derived organoids have been widely used to study the impact of extrinsic niche factors on stem cells. However, they are rarely used to study endogenous gene function due to the lack of efficient gene manipulation tools. Previously, we established a human foetal lung organoid system (Nikolić et al., 2017). Here, using this organoid system as an example we have systematically developed and optimised a complete genetic toolbox for use in tissue-derived organoids. This includes 'Organoid Easytag' our efficient workflow for targeting all types of gene loci through CRISPR-mediated homologous recombination followed by flow cytometry for enriching correctly-targeted cells. Our toolbox also incorporates conditional gene knock-down or overexpression using tightly-inducible CRISPR interference and CRISPR activation which is the first efficient application of these techniques to tissue-derived organoids. These tools will facilitate gene perturbation studies in tissue-derived organoids facilitating human disease modelling and providing a functional counterpart to many on-going descriptive studies, such as the Human Cell Atlas Project.
Keywords
Human, developmental biology
Sponsorship
Cancer Research UK (Core Support for Gurdon Institute C6946/A24843)
National Research Foundation of Korea (2018R1A6A3A03012122)
Cancer Research UK Core Support for Gurdon Institute (C6946/A24843)
Wellcome Trust PhD Studentship (109146/Z/15/Z)
Wellcome Trust PhD studentship (102175/B/13/Z)
Wellcome Trust Core Support for Gurdon Institute (203144/Z/16/Z)
Medical Research Council (MR/P009581/1, New Investigator Research Grant MR/T001917/1)
Medical Research Council New Investigator Research Grant (MR/T001917/1)
Wellcome Trust (Core Support for Gurdon Institute 203144/Z/16/Z, PhD studentship 102175/B/13/Z, PhD Studentship 109146/Z/15/Z)
Identifiers
PMC8553336, 34612202
External DOI: https://doi.org/10.7554/elife.67886
This record's URL: https://www.repository.cam.ac.uk/handle/1810/330412
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