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

  • Shoko Ishibashi
  • Nick R. Love
  • Enrique AmayaEmail author
Protocol
Part of the Methods in Molecular Biology book series (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.

Key words

I-SceI meganuclease Transgenesis Gateway 

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Copyright information

© Springer Science+Business Media, LLC 2012

Authors and Affiliations

  1. 1.The Healing Foundation Centre, The Faculty of Life SciencesUniversity of ManchesterManchesterEngland, UK
  2. 2.Healing Foundation Centre, Faculty of Life SciencesUniversity of ManchesterManchesterEngland, UK

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