Fig. 1: An orthogonal antibody eCLIP reveals transcriptome-wide Cas9-human RNA interactions.
From: Crosstalk between CRISPR-Cas9 and the human transcriptome
a Orthogonal V5/FLAG dSpCas9 eCLIP (enhanced eCLIP) experimental design. The experiment in transfected HEK 293T cells found 478 reproducible peaks across 381 human genes. All eCLIPs were performed in two bioreplicates per condition and were designed with two controls: size-matched inputs from dSpCas9 transfections; and antibody immunoprecipitations of empty vector transfections. b Immunofluorescent imaging of expressed V5/3xFLAG-dSpCas9 displays predominantly cytoplasmic cellular localization of dSpCas9 in HEK 293T cells. Experiments were performed independently in triplicate with similar results, with a representative image shown. c Highest enriched peak per gene log2(IP read count/size-matched input read count) enrichment score for V5 vs. FLAG eCLIPs. CDIP1(Cell Death Inducing p53 Target 1) is the top hit. d Gene regions of eCLIP peaks, with 3′ UTR the most represented.
