GSE123606
MOWChIP-seq for low-input and high-throughput profiling of genome-wide histone modifications
Epigenetic mechanisms such as histone modifications and DNA methylation form a critical layer of control that regulates gene activities. Histone modifications play critical roles in adaptive tuning of chromatin structures. Profiling various histone modifications at the genome scale using primary tissues from animal and human samples is an important step for functional studies of epigenomes and epigenomics-based precision medicine. Because the profile of a histone mark is highly specific to a particular cell type, isolation of a cell type of interest from primary tissues is often necessary to generate a homogeneous cell population and such operation tends to yield a low number of cells. In addition, high-throughput processing is desired in such effort due to the number of histone marks of interests and the potential volume of samples in a hospital setting. In this protocol, we describe detailed information on device fabrication, setup, and operation of microfluidic oscillatory washing-based chromatin immunoprecipitation followed by sequencing (MOWChIP-seq) for profiling histone modifications using as few as 30-100 cells per assay with a throughput as high as 8 assays in a run. The critical step of MOWChIP-seq operation involves flowing of chromatin fragments through a packed bed of antibody-coated beads followed by a vigorous microfluidic oscillatory washing. The ChIP process is semi-automated for reduced labor and improved reproducibility. We have used the protocol to study a number of histone modifications in various types of mouse and human tissue types ranging from isolated nuclei of brain cells to cell subtypes isolated from human breast tissues. Overall design: We examined genome-wide H3K9me3, H3K36me3, H3K79me2 profiles of nuclei isolated from mouse prefrontal cortex, 1000 nuclei per assay. The ChIP experiment was performed using half of 8-unit MOWChIP device, which performed 2 replicat experments for H3K9me3, H3K36me3 and H3K79me2 in every run under the same operating conditions.