Fig. 3. Characterization of factors affecting assay sensitivity and evaluation of assay specificity in eRNA detection.

a, Genome-wide distribution of sequencing reads originated from intergenic regions, introns, exons, and promoters detected by different assays. b, A genome browser snapshot of a gene (FAM89A) and its enhancer (highlighted in yellow), demonstrating the different patterns of signals captured by TSS- (enriched in the promoter and enhancer regions) vs. NT- (enriched in the promoter and gene body regions) assays. Signals are normalized by reads per million (RPM). c, Specificity in detecting eRNAs. Top track: differences of read coverage among CRISPR-identified enhancer ($n=803$) and non-enhancer ($n=\mathrm{6,777}$) loci, a pseudo-count (1) was added to each locus, and the coverage was log-transformed. Bottom track: signal-to-noise ratios depicted in forms of fold enrichment (FE), the results are based on bootstrapped samples ($n=\mathrm{10,000}$), median statistics are used to calculate the fold changes. In the box plot, the center dots, box limits, and whiskers denote the median, upper and lower quartiles, and 1.5× interquartile range, respectively. d, False discovery rates (FDRs) estimated by the overlap between the top 20,000 genomic bins and the reference (803 CRISPR-identified and 6,777 non-enhancer) loci. Downsampled libraries were used ($n=3$); values and error bars represent the mean and SD.