Comparison of morpholino based translational inhibition during the development of Xenopus laevis and Xenopus tropicalis.
Morpholino (MO) based inhibition of translational initiation represents an attractive methodology to eliminate gene function during Xenopus development (Heasman et al., 2000). However, the degree to which a given target protein can be eliminated and the longevity of this effect during embryogenesis has not been documented. To examine the efficacy of MOs, we have used transgenic Xenopus lines that harbour known numbers of integrations of a GFP reporter under the control of the ubiquitous and highly expressed CMV promoter (Fig. 1a). In addition we have investigated the longevity of the inhibitory effect by using transgenic lines expressing GFP specifically in the lens of tadpoles. These transgenic lines represent the ideal control for the technique as the promoters are highly expressed and GFP can be easily detected by fluorescence and immunoblotting. Moreover, as GFP has no function in development, the levels of inhibition can be tested in an otherwise normal individual. Here we report that MOs are able to efficiently and specifically inhibit the translation of GFP in transgenic lines from Xenopus laevis and Xenopus tropicalis and the inhibitory effect is long-lived, lasting into the tadpole stages. genesis 30:110--113, 2001.
Nutt
S L
SL
Wellcome/CRC Institute and Department of Zoology, University of Cambridge, Cambridge, United Kingdom.
Bronchain
O J
OJ
Hartley
K O
KO
Amaya
E
E
D000818:Animals; D030801:Animals, Genetically Modified; D003459:Crystallins / Q000235:genetics; D004625:Embryo, Nonmammalian / Q000196:embryology / Q000378:metabolism; D005260:Female; D018507:Gene Expression Regulation, Developmental; D017930:Genes, Reporter / Q000235:genetics; D049452:Green Fluorescent Proteins; D007814:Larva / Q000235:genetics / Q000254:growth & development; D008164:Luminescent Proteins / Q000235:genetics / Q000378:metabolism; D008297:Male; D009025:Morpholines / Q000378:metabolism*; D016376:Oligonucleotides, Antisense / Q000235:genetics / Q000378:metabolism*; D011401:Promoter Regions, Genetic / Q000235:genetics; D014176:Protein Biosynthesis; D014981:Xenopus / Q000196:embryology / Q000235:genetics* / Q000254:growth & development*; D014982:Xenopus laevis / Q000196:embryology / Q000235:genetics* / Q000254:growth & development*
D003459:Crystallins; D008164:Luminescent Proteins; D009025:Morpholines; D016376:Oligonucleotides, Antisense; D049452:Green Fluorescent Proteins; C037574:morpholine