Manipulation of Gene Function in Xenopus laevis

Abstract

Xenopus laevis embryos are particularly well suited to address questions requiring either knockdown or overexpression of genes in a tissue-specific fashion during vertebrate embryonic development. These manipulations are achieved by targeted injection of either antisense morpholino oligonucleotides, or synthetic mRNAs, respectively, into the early embryo. Herein we offer detailed protocols describing how to design and perform these experiments successfully, as well as a brief discussion of considerations for performing a microarray analysis in this organism.

The prodomain of BMP4 is necessary and sufficient to generate stable BMP4/7 heterodimers with enhanced bioactivity in vivo

“…Embryos were obtained, microinjected, and cultured as described (54). Embryonic stages are according to Nieuwkoop and Faber (55).…”

Fibronectin type III and intracellular domains of Toll-like receptor 4 interactor with leucine-rich repeats (Tril) are required for developmental signaling

“…Xenopus embryo culture and manipulation-Animal procedures followed protocols approved by the University of Utah Institutional Animal Care and Use Committee. Embryos were obtained, microinjected, and cultured as described (26). Embryos were processed for in situ hybridization as described (27) except that the vitelline coat was not removed prior to fixation and BM purple (Roche) was used as a substrate.…”

Centriole biogenesis and function in multiciliated cells

“…One significant advantage of using Xenopus as a model organism is the ease of inducing localized protein expression via direct injection of in vitro synthesized messenger RNA (mRNA) without the requirement of genome-level manipulations (Mimoto & Christian, 2011; Sive et al, 2007e, 2010a; Sive, Grainger, & Harland, 2010b). The detailed protocols for manipulating early Xenopus embryos using mRNA injection method has been extensively described and profiled.…”

“…While these recent studies have begun to provide a better picture on this structure, clearly there is still much to be learned. Many of the techniques used to study centriole biology in Xenopus rely on standard approaches that have been described in detail elsewhere (Mimoto & Christian, 2011;Sive, Grainger, & Harland, 1998, 2007c, 2007e, 2010a; Werner & Mitchell, 2013). Here, we aim to highlight specific tools, reagents, and experimental conditions that will facilitate the study of centriole amplification in MCCs.…”