Dynamic visualization of transcription and RNA subcellular localization in zebrafish

摘要: Live imaging of transcription and RNA dynamics has been successful in cultured cells tissues vertebrates but is challenging to accomplish vivo. The zebrafish offers important advantages study these processes – optical transparency during embryogenesis, genetic tractability rapid development. Therefore, an intact vertebrate organism, we have adapted the MS2 RNA-labeling system zebrafish. By using this binary coexpress a fluorescent bacteriophage coat protein (MCP) interest tagged with multiple copies hairpin MS2-binding site (MBS), live-cell at single molecule resolution achieved other organisms. Here, Gateway-compatible labeling system, generated stable transgenic lines expressing MCP, validated MBS-MCP interaction applied investigate zygotic genome activation (ZGA) localization primordial germ (PGCs) Although cleavage stage are initially transcriptionally silent, detect MS2-tagged transcripts driven by βactin promoter ∼3-3.5 h post-fertilization, consistent previously reported ZGA. Furthermore, show that nanos3 3′UTR localize PGCs, where they diffusely cytoplasmic within larger accumulations reminiscent those displayed endogenous nanos3. These tools provide new avenue for molecules vertebrate. Together techniques targeted editing, will be valuable tool tag RNAs developmental processes.