Generation of trangenic Xenopus laevis using the Sleeping Beauty transposon system.
Using the Sleeping Beauty (SB) transposon system, we have developed a simple method for the generation of Xenopus laevis transgenic lines. The transgenesis protocol is based on the co-injection of the SB transposase mRNA and a GFP-reporter transposon into one-cell stage embryos. Transposase-dependent reporter gene expression was observed in cell clones and in hemi-transgenic animals. We determined an optimal ratio of transposase mRNA versus transposon-carrying plasmid DNA that enhanced the proportion of hemi-transgenic tadpoles. The transgene is integrated into the genome and may be transmitted to the F1 offspring depending on the germline mosaicism. Although the transposase is necessary for efficient generation of transgenic Xenopus, the integration of the transgene occurred by an non-canonical transposition process. This was observed for two transgenic lines analysed. The transposon-based technique leads to a high transgenesis rate and is simple to handle. For these reasons, it could present an attractive alternative to the classical Restriction Enzyme Mediated Integration (REMI) procedure.
Sinzelle
L
L
Transgenèse et Génétique des Amphibiens, CNRS UMR 8080, IBAIC, Université Paris-Sud, 91405, Orsay Cedex, France.
Vallin
J
J
Coen
L
L
Chesneau
A
A
Du Pasquier
D
D
Pollet
N
N
Demeneix
B
B
Mazabraud
A
A
D000818:Animals; D030801:Animals, Genetically Modified; D004251:DNA Transposable Elements; D004625:Embryo, Nonmammalian; D018014:Gene Transfer Techniques; D049452:Green Fluorescent Proteins / Q000235:genetics; D008845:Microinjections; D012333:RNA, Messenger / Q000008:administration & dosage; D019076:Transgenes; D019895:Transposases / Q000008:administration & dosage* / Q000235:genetics; D014982:Xenopus laevis
D004251:DNA Transposable Elements; D012333:RNA, Messenger; D049452:Green Fluorescent Proteins; D019895:Transposases