Abstract
Efficient and reliable methods for gene editing are critical for the generation of loss-of-gene function stem cells and genetically modified mice. Here, we outline the application of CRISPR-Cas9 technology for gene editing in mouse embryonic stem cells (mESCs) to generate knockout ESC chimeras for the fast-tracked analysis of gene function. Furthermore, we describe the application of gene editing directly to mouse epiblast stem cells (mEpiSCs) for modelling germ layer differentiation in vitro.
This is a preview of subscription content, log in via an institution.
Buying options
Tax calculation will be finalised at checkout
Purchases are for personal use only
Learn about institutional subscriptionsReferences
Urnov FD, Rebar EJ, Holmes MC, Zhang HS, Gregory PD (2010) Genome editing with engineered zinc finger nucleases. Nat Rev Genet 11:636–646
Hsu PD, Zhang F (2012) Dissecting neural function using targeted genome engineering technologies. ACS Chem Neurosci 3:603–610
Ran FA, Hsu PD, Wright J, Agarwala V, Scott DA, Zhang F (2013) Genome engineering using the CRISPR-Cas9 system. Nat Protoc 8:2281–2308
Osteil P, Studdert J, Wilkie E, Fossat N, Tam PP (2016) Generation of genome-edited mouse epiblast stem cells via a detour through ES cell-chimeras. Differentiation 91:119–125
Fossat N, Ip CK, Jones VJ, Studdert JB, Khoo PL et al (2015) Context-specific function of the LIM homeobox 1 transcription factor in head formation of the mouse embryo. Development 142:2069–2079
Brinkman EK, Chen T, Amendola M, van Steensel B (2014) Easy quantitative assessment of genome editing by sequence trace decomposition. Nucleic Acids Res 42:e168
Edgar RC (2004) MUSCLE: multiple sequence alignment with high accuracy and high throughput. Nucleic Acids Res 32:1792–1797
Edgar RC (2004) MUSCLE: a multiple sequence alignment method with reduced time and space complexity. BMC Bioinformatics 5:113
Acknowledgments
Our work was supported by grants from the Australian Research Council (DP 160103651, DP 160100933), the National Health and Medical Research Council of Australia (1127976), and the late Mr. James Fairfax (Bridgestar Foundation). PPLT is a NHMRC Senior Principal Research Fellow (Grant 1110751).
Author information
Authors and Affiliations
Corresponding author
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2019 Springer Science+Business Media, LLC, part of Springer Nature
About this protocol
Cite this protocol
Sibbritt, T. et al. (2019). Gene Editing of Mouse Embryonic and Epiblast Stem Cells. In: Bertoncello, I. (eds) Mouse Cell Culture. Methods in Molecular Biology, vol 1940. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-9086-3_6
Download citation
DOI: https://doi.org/10.1007/978-1-4939-9086-3_6
Published:
Publisher Name: Humana Press, New York, NY
Print ISBN: 978-1-4939-9085-6
Online ISBN: 978-1-4939-9086-3
eBook Packages: Springer Protocols