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Microinjection and Oviduct Transfer Procedures for Rat Model Generation with CRISPR-Cas9 Technology

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Microinjection

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1874))

Abstract

Since the first knockout rat model was generated with zinc-finger nucleases (ZFNs) by Geurt’s group in 2009, the demand for making targeted rat models has increased tremendously. The advent of the clustered regularly interspaced short palindromic repeats-CRISPR associated protein 9 (CRISPR-Cas9) system provides researchers with a more efficient method for producing modified animals, which has since then been developed and applied in rat. Since we established a rat model production system at our facility in 2014, we have consistently generated rat models. Due to differences in physiology and embryology between mouse and rat, species-specific protocols for superovulation conditions, microinjection, and embryo transfer (among others) are required. There are over 100 rat strains, and Sprague Dawley is one of the commonly used outbred strains in biomedical research. In this chapter, we describe in detail a range of topics including donor and recipient preparation, microinjection setup, CRISPR reagent preparation, and oviduct transfer procedures for making rat models in the Sprague Dawley background.

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Acknowledgments

We thank the Technology Development Group, Genotyping Analysis Lab, Colony Resource Group, Animal Resource Group, and Transgenic Technology Co-Op group at Genentech for their collaboration and support for producing rat models. We would also like to thank Dr. Juan M. Reyes for his suggestions and assistance with editing this chapter, and Natasha O’Neil for photography.

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Authors and Affiliations

  1. Embryo Technology Group, gRED Animal Resources, Genentech, Inc., South San Francisco, CA, USA

    Tuija M. Alcantar & Xin Y. Rairdan

Authors
  1. Tuija M. Alcantar
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  2. Xin Y. Rairdan
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Corresponding author

Correspondence to Xin Y. Rairdan .

Editor information

Editors and Affiliations

  1. National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, MD, USA

    Chengyu Liu

  2. National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, MD, USA

    Yubin Du

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Alcantar, T.M., Rairdan, X.Y. (2019). Microinjection and Oviduct Transfer Procedures for Rat Model Generation with CRISPR-Cas9 Technology. In: Liu, C., Du, Y. (eds) Microinjection. Methods in Molecular Biology, vol 1874. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-8831-0_16

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  • DOI: https://doi.org/10.1007/978-1-4939-8831-0_16

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  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-8830-3

  • Online ISBN: 978-1-4939-8831-0

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