Fig. 2: Enhancer targeting with truncated guides induces EPB41 repression.
From: Multi-locus CRISPRi targeting with a single truncated guide RNA
a Schematic of EPB41 genomic regulatory region and full-length/truncated guides targeting each of 4 TF motifs. b TF motifs (JASPAR) targeted by the guides with protospacer adjacent motif (PAM) orientation denoted. c Number of sequence match target sites in the genome (hg38) for the indicated motif directed guide. d Quantitative real time PCR analysis of EPB41 expression in K562 cells with respective guide CRISPRi treatment conditions. Safe (Safe Harbor-targeting) guide = g[20nt], promoter guide = 21nt, enh1 = g[20nt], enh2 = 21nt, enh3 = g[20nt] (enhancer guides from Gasperini et al. 2019). Data are mean +/− SEM. e Violin plot summary of EPB41 expression data in panel d organized by guide length or targeting region. TF-targeting guides (n = 16; 4 guides per spacer length), sgEnh (n = 12; 3 guides), and Promoter (n = 4; 1 guide, all points shown) are the product of 2 biological replicates per guide run in duplicate. Safe is the sum of 3 biological replicates run 2-3 times each (n = 8). Data are normalized to the mean CT value of Safe Harbor actin-b control probe. Results are analyzed by one-way ANOVA. ****P < 0.0001 for all guide lengths compared with Safe Harbor guide. P = 8.24 × 10−7, 9.95 × 10−7, 5.62 × 10−4, 3.98 × 10−5, and 0.01 for Safe vs. 21nt,g[20nt]; 14nt,g[13nt]; 11nt; sgEnh; and Promoter respectively. Source data are provided as a Source Data file.
