Abstract
Molecular genetic strategies applying embryonic stem cell (ES cell) technologies to study the function of a gene in mice or to generate a mouse model for a human disease are continuously under development. Next to (conditional) inactivation of genes the application and importance of approaches to generate knock-in mutations are increasing. In this chapter the principle and application of recombinase-mediated cassette exchange (RMCE) are discussed as being a new emerging knock-in strategy, which enables easy generation of a series of different knock-in mutations within one gene. An RMCE protocol, which was used to generate a series of different knock-in mutations in the Lrp1 gene of ES cells, is described in detail as an example of how RMCE can be used to generate highly efficiently an allelic series of differently modified ES cell clones from a parental modified ES cell clone. Subsequently the differently modified ES cell clones can be used to generate an allelic series of mutant knock-in mice.
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Acknowledgement
This work was supported by grants from the Fund for Scientific Research-Flanders (Fonds voor Wetenschappelijk Onderzoek Vlaanderen, FWO-Vlaanderen G.0529.08, G.0689.10, and G.0830.11) and the Concerted Actions Program of the KU Leuven (GOA/12/016) to Anton Roebroek. The work was also funded by a PhD grant of the Agency for Innovation by Science and Technology (IWT) to Bart Van Gool.
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Roebroek, A.J.M., Van Gool, B. (2014). Generation of an Allelic Series of Knock-In Mice Using Recombinase-Mediated Cassette Exchange (RMCE). In: Singh, S., Coppola, V. (eds) Mouse Genetics. Methods in Molecular Biology, vol 1194. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-1215-5_4
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DOI: https://doi.org/10.1007/978-1-4939-1215-5_4
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