High-throughput transgenesis in Xenopus using I-SceI meganuclease.
In this report we describe an easy, highly efficient transgenesis method for Xenopus. The method is very simple; a commercially available meganuclease, I-SceI, is incubated with a transgene construct carrying its recognition sites, and is subsequently microinjected into fertilized eggs. Approximately 30% (in Xenopus tropicalis) or 20% (in Xenopus laevis) of injected embryos exhibit non-mosaic, promoter-dependent transgene expression, and transgenes from the founder animals are transmitted to offspring. The method is compatible with mRNA or antisense morpholino oligonucleotide injection, and these secondary reagents can be introduced simultaneously or sequentially with a transgene to test their interaction. This high-throughput transgenic technique will be a powerful tool for studying the complex wiring of regulatory networks at the genome-wide level, as well as for facilitating genetic studies in the rapidly breeding diploid frog, X. tropicalis.
Ogino
Hajime
H
Department of Biology, University of Virginia, Charlottesville, Virginia 22904, USA.
McConnell
William B
WB
Grainger
Robert M
RM
D000818:Animals; D015252:Deoxyribonucleases, Type II Site-Specific; D018014:Gene Transfer Techniques; D008845:Microinjections / Q000379:methods; D029701:Saccharomyces cerevisiae Proteins; D014981:Xenopus / Q000196:embryology / Q000235:genetics*
D029701:Saccharomyces cerevisiae Proteins; C037725:SCEI protein, S cerevisiae; D015252:Deoxyribonucleases, Type II Site-Specific