A gene trap approach in Xenopus.
The frog transgenesis technique ultimately promises to make mutagenesis possible through random insertion of plasmid DNA into the genome. This study was undertaken to evaluate whether a gene trap approach combined with transgenesis would be appropriate for performing insertional mutagenesis in Xenopus embryos. Firstly, we confirmed that the transgenic technique results in stable integration into the genome and that transmission through the germline occurs in the expected Mendelian fashion. Secondly, we developed several gene trap vectors, using the green fluorescent protein (GFP) as a marker. Using these vectors, we trapped several genes in Xenopus laevis that are expressed in a spatially restricted manner, including expression in the epiphysis, the olfactory bulb and placodes, the eyes, ear, brain, muscles, tail and intestine. Finally, we cloned one of the trapped genes using 5' rapid amplification of cDNA ends polymerase chain reaction (RACE PCR). These results suggest that the transgenic technique combined with a gene trap approach might provide a powerful method for generating mutations in endogenous genes in Xenopus.
Bronchain
O J
OJ
Wellcome/CRC Institute, Department of Zoology, University of Cambridge, Tennis Court Road, Cambridge, CB2 1QR, UK.
Hartley
K O
KO
Amaya
E
E
D000595:Amino Acid Sequence; D000818:Animals; D030801:Animals, Genetically Modified; D001483:Base Sequence; D003459:Crystallins / Q000235:genetics; D004274:DNA, Recombinant / Q000235:genetics; D015870:Gene Expression; D005821:Genetic Techniques; D005822:Genetic Vectors; D049452:Green Fluorescent Proteins; D008164:Luminescent Proteins / Q000235:genetics; D008969:Molecular Sequence Data; D016254:Mutagenesis, Insertional; D016133:Polymerase Chain Reaction / Q000379:methods; D011993:Recombinant Fusion Proteins / Q000235:genetics; D014981:Xenopus / Q000196:embryology / Q000235:genetics*
D003459:Crystallins; D004274:DNA, Recombinant; D008164:Luminescent Proteins; D011993:Recombinant Fusion Proteins; D049452:Green Fluorescent Proteins