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A Simple Method of Transgenesis Using I-Sce I Meganuclease in Xenopus

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Xenopus Protocols

Part of the book series: Methods in Molecular Biology ((MIMB,volume 917))

Abstract

Here we present a protocol for generating transgenic embryos in Xenopus using I-SceI meganuclease. This method relies on integration of DNA constructs, containing one or two I-SceI meganuclease sites. It is a simpler method than the REMI method of transgenesis, and it is ideally suited for generating transgenic lines in Xenopus laevis and Xenopus tropicalis. In addition to it being simpler than the REMI method, this protocol also results in single copy integration events rather than tandem concatemers. Although the protocol we describe is for X. tropicalis, the method can also be used to generate transgenic lines in X. laevis. We also describe a convenient method for designing and generating complex constructs for transgenesis, named pTransgenesis, based on the Multisite Gateway® cloning, which include I-SceI sites and Tol2 elements to facilitate genome integration.

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Author information

Authors and Affiliations

  1. The Healing Foundation Centre, The Faculty of Life Sciences, University of Manchester, Manchester, England, UK

    Shoko Ishibashi & Nick R. Love

  2. Healing Foundation Centre, Faculty of Life Sciences, University of Manchester, Manchester, England, UK

    Enrique Amaya

Authors
  1. Shoko Ishibashi
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  2. Nick R. Love
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  3. Enrique Amaya
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Corresponding author

Correspondence to Enrique Amaya .

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Editors and Affiliations

  1. , Institute of Medical Sciences, University of Aberdeen, FORESTERHILL, ABERDEEN, AB25 2ZD, United Kingdom

    STEFAN HOPPLER

  2. , Department of Biological Science, University of Calgary, University Drive NW 2500, Calgary, T2N 1N4, Canada

    Peter D Vize

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Ishibashi, S., Love, N.R., Amaya, E. (2012). A Simple Method of Transgenesis Using I-Sce I Meganuclease in Xenopus . In: HOPPLER, S., Vize, P. (eds) Xenopus Protocols. Methods in Molecular Biology, vol 917. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-61779-992-1_12

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  • DOI: https://doi.org/10.1007/978-1-61779-992-1_12

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  • Publisher Name: Humana Press, Totowa, NJ

  • Print ISBN: 978-1-61779-991-4

  • Online ISBN: 978-1-61779-992-1

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