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1) Product Images from "PAtCh-Cap: input strategy for improving analysis of ChIP-exo data sets and beyond"
Article Title: PAtCh-Cap: input strategy for improving analysis of ChIP-exo data sets and beyond
Journal: Nucleic Acids Research
Figure Legend Snippet: Application of PAtCh-Cap to CTCF ChIP-exo data allowed for significant artifact removal and improved confidence in peak identification. ( A ) To identify high-confidence CTCF peaks, all peaks called with a 0.05 q -value threshold from the ChIP-exo data with (red) and without (black) input treatment were plotted as the –log(q-value) versus ranked peak number. High confidence peaks were determined to be those characterized by a –log( q -value) higher than the inflection point and a line with a slope of –tan 1 to the curve (denoted by vertical lines). ( B ) Venn diagram demonstrating the overlap of high-confidence peaks identified from data sets with and without input treatment (top). The number of CTCF motifs found within each pool is denoted and clearly shows that the percentage of CTCF containing peaks relative to the total increases substantially after input treatment. Venn diagrams for the overlap of blacklisted peaks with each of the above regions (bottom).
Techniques Used: Chromatin Immunoprecipitation
Figure Legend Snippet: Representative CTCF ChIP-exo read coverage tracks for the pericentromeric region of chromosome 1 ( A ) and the promoter of the KCNJ3 gene ( B ). The CTCF reads (blue) were normalized to the reads from the input control (green) using MACS2 ( 27 ) to generate the enrichment read coverage tracks (red). Peaks identified by the MACS2 peak caller (represented in the .bed tracks) are denoted as red or blue vertical lines for the CTCF ChIP-exo data sets with and without input treatment, respectively. Analysis of the genomic sequences underneath the remaining peaks after input treatment (vertical red lines) definitively showed that these sites contain the core CTCF motif as evidenced by alignment of the CTCF sequence logo beneath.
Techniques Used: Chromatin Immunoprecipitation, Genomic Sequencing, Sequencing
Figure Legend Snippet: From the input treated CTCF ChIP-exo data set, ( A ) read tag distributions around all genomic CTCF-bound sites shown in the four binned motif combinations (right panel) were centered on the midpoint of the CTCF consensus to generate a heat map (top left) which is summed below as an aggregate plot. Denoted in blue and red are the sense and antisense strand read enrichments around the core CTCF motif, respectively. The centralized CTCF core sequence and adjacent motifs are depicted above a color map representation of 50 bp DNA stretches containing the various motif combinations (right panel). ( B ) Heat maps from RNA-seq data depicting gene transcripts exhibiting a two-fold up- (green) or down-regulation (red) after CTCF depletion relative to the scrambled siRNA control (Scr). For each motif group, CTCF promoter occupation sites (defined as ±1000 bps around the transcription start site (TSS)) were intersected with the RNA-seq data and resulting altered gene sets were binned as individual heat maps. ( C ) Each gene set from ( B ) was subjected to Ingenuity Pathway Analysis (IPA, www.ingenuity.com ) to identify biological pathways uniquely modulated by each of the CTCF motif combinations. ( D–F ) The same analyses in ( A–C ) were performed separately on the core CTCF consensus with the newly identified 3′-CTCF motif.
Techniques Used: Chromatin Immunoprecipitation, Sequencing, RNA Sequencing Assay, Indirect Immunoperoxidase Assay
Article Snippet: Once covalently bound to the magnetic beads, the input samples were treated identically as described above for the CTCF ChIP-exo samples utilizing reagents and materials from the