J Vis Exp 2026 Mar 27;229:. doi: 10.3791/70591.

Metabolic Labeling of the Nascent Transcriptome in Xenopus Early Embryogenesis.

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Early embryogenesis requires new transcription of zygotic genes from the initially dormant zygotic genome after fertilization. A major challenge has been to accurately detect the new transcripts and determine their time of expression during zygotic genome activation (ZGA). Here, we provide detailed procedures for metabolic labeling of the nascent transcripts by 5-ethynyl-uridine (5-EU) in early Xenopus laevis embryos. After fertilization, 5-EU is microinjected into the 1-cell stage embryos to metabolically label the newly transcribed RNAs during ZGA. The nascent EU-RNAs are subsequently purified after biotinylation via click chemistry-mediated reactions, and the nascent transcriptome is determined using next-generation sequencing. Importantly, this method has high sensitivity and specificity that allow the characterization of lowly activated genes. Thus, this approach provides a powerful tool to study the dynamics of gene transcription during early development, and it can be generalized to other embryonic systems or tissues.

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