Fig. 1

Identification and initial screening for regulatory elements active in E5 chick retinae. (a) Generation of ATAC-seq libraries of ThrbCRM1-positive and ThrbCRM1-negative cell populations. Chick retinae at embryonic day 5 (E5) were electroporated ex vivo with both ThrbCRM1::GFP and UbiqC::TdT plasmids and incubated in culture for 18–22 h prior to dissociation. Dissociated cells were sorted via FACS into GFP and TdT double-positive cells, and GFP-negative, TdT-positive cells. Each population was processed for ATAC-seq. (b) Visualization of aligned ATAC-seq reads to the galGal5 genome in UCSC Genome Browser in intergenic region between Onecut1 and WDR72 (labelled). Active regulatory elements represented by colored, labelled lines based on activity level from alkaline phosphatase assay (c) Alkaline phosphatase reporter assay to screen for regulatory activity. E5 chick retinae were electroporated with CRE::AP plasmids and CAG::mCherry plasmids. Empty Stagia3 vector (No Enhancer) represents the negative control. Dotted boxed region corresponds to magnified inset shown in bottom right corner. Scale bars in the first panel and inset represent 500 μm and apply to all panels