Fig. 1: The RNA-binding activity of HARD protein.
From: Capture of RNA-binding proteins across mouse tissues using HARD-AP
a Schematic diagram of HARD protein. HARD protein is composed of an SSB OB fold domain, a LINE-1 ORF1p C-1/3 domain and a flexible linker. b The Coomassie blue stained SDS-PAGE gel showing the purified recombinant EGFP-HARD and EGFP. This experiment was repeated once with similar results. c Isothermal titration calorimetry assay showing the binding of EGFP-HARD to single-stranded RNA (ssRNA) and double-stranded RNA (dsRNA). Shown are normalized data with the best fits (solid lines). Raw data are shown in Supplementary Fig. 1f. d Percentage of input RNA species isolated using HARD beads and EGFP beads as determined using qRT-PCR. Data are means ± SD; three independent biological samples were used for the analysis (n = 3); significance was determined using the two-tailed Student’s t-test. e Scatter plot showing the correlation of normalized RNA-seq signals (FPKM) of HARD beads-enriched RNAs (HARD-AP) and input RNAs (n = 60,623). Two independent biological samples are labeled as rep1 and rep2. The colors scale indicates dot density. The Pearson correlation coefficients are given. f Volcano plot showing distributions of RNAs differentially detected in HARD-AP and Input samples (n = 60,623). Significantly differentially detected RNAs are labeled with red dots. The significance (p) was determined using the two-tailed Student’s t-test and further adjusted using the Benjamini-Hochberg correction for multiple testing (p-adjust). g Metagene representation of RNA-seq signals of HARD-AP and Input samples in bodies of genes (n = 60,623). h Percentage of indicated RNA species and RNA-seq read distributions along genes in HARD-AP and input samples. i Genome browser tracks showing normalized RNA-seq signals along indicated genomic loci in HARD-AP and input samples. Source data for (b, e-f) are provided as a Source Data file.
