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).
Siprashvili et al. BMC Genomics 2012 13:633 doi:10.1186/1471-2164-13-633