XB-ART-48095Methods April 1, 2014; 66 (3): 422-32.
Design and use of transgenic reporter strains for detecting activity of signaling pathways in Xenopus.
Embryos and larvae of vertebrate species with external development are ideal subjects for investigating the dynamic spatiotemporal activity of developmental signaling pathways. The availability of efficient transgene technologies in Xenopus and zebrafish and the translucency and/or transparency of their embryos and larvae make these two species attractive for direct in vivo imaging of reporter gene expression. In this article we describe the design of efficient signaling reporters, using the Wnt/β-catenin pathway as a representative example. We define methods for validating the reporter constructs and describe how they can be used to generate stable transgenic lines in Xenopus. We provide efficient methods used in our laboratory for raising the tadpoles and froglets rapidly to sexual maturity. We further discuss how the reporter lines can be used for delineating the dynamic activity of a signaling pathway and how modulators of the pathway can be scrutinized via chemical intervention and the micro-injection of synthetic RNAs or morpholinos. The strategic outline discussed in this paper provides a template for studying other developmental signaling pathways in Xenopus.
PubMed ID: 23816788
Article link: Methods
Species referenced: Xenopus
Genes referenced: ctnnb1 lef1
Morpholinos: lef1 MO5