PIC-RNA-seq for Region-Specific Transcriptomic Analysis of Chicken Limb Bud Progenitors

Region-specific RNA sequencing is a powerful approach for investigating tissue differentiation and dynamic changes in gene expression during embryonic development. The chicken embryo has long served as an important model system in developmental biology. However, the limited availability of tissue-specific reporter lines makes region-specific RNA-seq approaches particularly valuable in this organism. Here, we applied photo-isolation chemistry-based RNA sequencing (PIC-RNA-seq) to the somatic lateral plate mesoderm (sLPM) before the emergence of limb bud progenitor cells (LPCs) and to early LPCs in chicken embryos. These analyses revealed the upregulation of multiple genes, including Hox genes, in LPCs, suggesting the initiation of their regional patterning program. This workflow enables visualization of dynamic changes in the gene expression profile of LPCs and should also be applicable to other tissues in avian embryos. Key features • An optimized PIC-RNA-seq protocol for the somatic lateral plate mesoderm (sLPM) and limb bud progenitor cells (LPCs) in chicken embryos. • Applicable to chicken embryos without the need for tissue-specific transgenic reporter lines. • Robust generation of high-quality RNA-seq datasets with sufficient sequencing depth, unique molecular identifiers (UMIs), and detected genes for downstream analyses. • Enables direct comparison between pre-specified sLPM and specified LPCs, allowing the detection of transcriptional changes accompanying early LPC specification.