Optimized ChIP-exo for mammalian cells and patterned sequencing flow cells
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By combining chromatin immunoprecipitation (ChIP) with an exonuclease digestion of protein-bound DNA fragments, ChIP-exo characterizes genome-wide protein-DNA interactions at near base-pair resolution. However, the widespread adoption of ChIP-exo has been hindered by several technical challenges, including lengthy protocols, the need for multiple custom reactions, and incompatibilities with recent Illumina sequencing platforms. To address these barriers, we systematically optimized and adapted the ChIP-exo library construction protocol for the unique requirements of mammalian cells and current sequencing technologies. We introduce a Mammalian-Optimized ChIP-exo (MO-ChIP-exo) protocol that builds upon previous ChIP-exo protocols with systematic optimization of crosslinking, harvesting, and library construction. We validate MO-ChIP-exo by comparing it to previously published ChIP-exo protocols and demonstrate its adaptability to both suspension (K562) and adherent (HepG2, mESC) cell lines. This improved protocol provides a more robust and efficient method for generating high-quality ChIP-exo libraries from mammalian cells.
SUMMARY
ChIP-exo is a genome-wide protein-DNA binding assay with unrivalled resolution, but its widespread adoption has been hindered by technical challenges, particularly when applied to mammalian cells or when used with recent sequencing platforms. We introduce a Mammalian-Optimized ChIP-exo (MO-ChIP-exo) protocol with key modifications that overcome previous technical hurdles. We demonstrate that our optimized protocol produces high-quality data comparable to previously published protocols and is adaptable for use with both suspension (K562) and adherent (HepG2, mESC) cell lines.
