MOBE-ChIP: A large-scale chromatin immunoprecipitation assay for cell type-specific studies

Abstract

Cell type-specific transcriptional regulators play critical roles in the generation and maintenance of multicellularity. As they are often expressed at low levels, in vivo DNA-binding studies of these regulators by standard chromatin immunoprecipitation (ChIP) assays are technically challenging. We describe here an optimized ChIP protocol termed Maximized Objects for Better Enrichment (MOBE)-ChIP, which enhances the sensitivity of ChIP assays for detecting cell type-specific signals. The protocol, which is based on the disproportional increase of target signals over background at higher scales, uses substantially greater volume of starting materials than conventional ChIPs to achieve high signal enrichment. This technique can capture weak binding events that are ambiguous in standard ChIP assays, and is useful both in gene-specific and whole-genome analysis. This protocol has been optimized for Arabidopsis, but should be applicable to other model systems with minor modifications. The full procedure can be completed within 3 days. Significance Statement In vivo DNA binding studies of cell type-specific transcriptional regulators by standard chromatin immunoprecipitation (ChIP) assays are technically challenging. Here we present a detailed protocol for an optimized ChIP procedure that significantly improves signal enrichment, allowing detection of cell type-specific signals and high quality genome-wide binding data.