Transcription factories have been characterized in cultured mammalian cells, but little is known about the regulation of these nuclear structures in different primary cell types. Using marine medaka, we observed transcription sites labeled by the metabolic incorporation of 5-fluorouridine (5-FU) into nascent RNA. Medaka was permeable to 5-FU in ambient water and became fully labeled within 4 hr of incubation. The incorporation of 5-FU was inhibited by the transcription inhibitor actinomycin D. The 5-FU incorporation sites were detected in the cell nucleus, and could be abolished by RNase digestion. The tissue distribution of 5-FU incorporation was visualized by immunocytochemistry on whole-mount specimens and histological sections. The 5-FU labeling appeared highly cell type specific, suggesting a regulation of the overall transcription activities at tissue level. Mapping of transcription factories by 5-FU incorporation in fish provides a useful and physiologically relevant model for studying the control of gene expression in the context of the functional organization of the cell nucleus. This manuscript contains online supplemental material at http://www.jhc.org. Please visit this article online to view these materials.