Chemical mapping of medaka and human CR4/5 domains. (A) Cartoon schematic of general RNA construct design, including the RNA sequence of interest flanked by unstructured RNA buffer sequences, a normalization RNA hairpin, and a reverse transcriptase priming site. (B) Lowest energy predicted secondary structure of medaka (left) and human (right) CR4/5 domain using RNAstructure. (C) (left) Chemical mapping of the medaka CR4/5 domain by SHAPE (1M7 probing) at 1 mM MgCl2. (D) Chemical mapping of the human CR4/5 domain by SHAPE (1M7 probing) at 1 mM MgCl2. For both (C,D), color coding in the bar plot and structure schematic is as described in C. Plotted normalized reactivity values are color-coded (red >0.7, yellow 0.3–0.7, and black <0.3). Each bar plotted represents experiments conducted in triplicate or greater with the respective standard deviation as error bars (right). Color-coded schematic of the reactivity data is shown on the RNAstructure predicted secondary structure.
