Figure 1

RNA incubation on human protein microarrays. (A) Experimental and analytical workflow. (B) RNA incubation signal on protein microarray. Sense [red] and antisense [green] strand signal is shown for the SOX2OT non-coding RNA, with pseudocolor images of independently probed arrays. Panels at left show the entire microarray spotted with ~9400 recombinant human proteins; the middle panel is an enlarged 484 protein spot sub-array and the right panels represent an enlargement of the binding signal demonstrating strand-specific binding to the RBPMS protein [GenBank:BC003608] [all proteins spotted in duplicate; sub-array positive controls boxed in red]. (C) Scatter plot of signal intensity above background for all proteins between two independent replicates of the HRAS sense mRNA. Pearson correlation r² value is shown at right. (D) RNA binding signal intensity over background distribution for TP53 sense RNA to the all proteins. Mean and Z-Scores (standard deviations from mean) are depicted in blue with selected fold change cutoff of 2.5 in red. A Z-Score ≥ 3 and signal intensity over background ≥ 2.5 are used to select significant RNA-protein binding event. (E) Venn diagram of significant hits from two independent HRAS mRNA incubations to protein array from (C), (p value, Fisher’s exact test).