Generating transgenic frog embryos by Restriction Enzyme Mediated Integration (REMI)

Shoko Ishibashi, Kristen L. Kroll, Enrique Amaya

Research output: Chapter in Book/Report/Conference proceedingChapterpeer-review

11 Scopus citations

Abstract

Here we present a protocol for generating transgenic embryos in Xenopus laevis and Xenopus tropicalis. The method includes three steps: (1) The preparation of high-speed egg extracts, which facilitates the replacement of protamines in sperm nuclei with nucleosomes and decondenses the chromatin of sperm nuclei; (2) The isolation of sperm nuclei; and (3) The mixing of sperm nuclei, restriction enzyme, and high-speed extract in vitro, following by nuclear transplantation into unfertilized eggs to generate the transgenic embryos. This procedure generates non-mosaic transgenic embryos at high frequency and efficiency.

Original languageEnglish
Title of host publicationXenopus Protocols
Subtitle of host publicationPost-Genomic Approaches
PublisherHumana Press Inc.
Pages185-203
Number of pages19
ISBN (Print)9781617799914
DOIs
StatePublished - 2012

Publication series

NameMethods in Molecular Biology
Volume917
ISSN (Print)1064-3745

Keywords

  • Restriction enzyme mediated integration
  • Transgenesis
  • Xenopus

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