mRNA Microinjection for Gene Expression Studies in Xenopus laevis and Xenopus tropicalis.

Microinjection of in vitro-transcribed, capped mRNA into Xenopus laevis and Xenopus tropicalis embryos provides a rapid and powerful approach for gene expression studies in vertebrate development. Owing to the large size, external development, and manipulability of Xenopus embryos, mRNA injection allows precise temporal and spatial control of expression and the ability to test both wild-type and mutant variants in vivo. Classic experiments using synthetic mRNAs identified key regulators of embryonic patterning and signaling, and the approach continues to underpin modern functional genomics and disease modeling. Integration of mRNA injection with CRISPR/Cas9-mediated knockouts, fluorescent reporters, and human variant testing has further enhanced its versatility. This chapter details practical methods for mRNA synthesis, quality control, and embryo injection, providing a reliable framework for controlled gene expression analysis and validation in Xenopus embryos.