MOWChIP-seq for low-input and multiplexed profiling of genome-wide histone modifications
| Autor: | Qiang Zhang, Chang Lu, Lynette B. Naler, Bohan Zhu, Yuan-Pang Hsieh, Travis W. Murphy |
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| Rok vydání: | 2019 |
| Předmět: |
Epigenomics
Chromatin Immunoprecipitation Cell type Sequence analysis Computer science Microfluidics Population Computational biology Genome Article General Biochemistry Genetics and Molecular Biology Epigenesis Genetic Histones 03 medical and health sciences 0302 clinical medicine Animals Humans Epigenetics education 030304 developmental biology 0303 health sciences education.field_of_study biology High-Throughput Nucleotide Sequencing Sequence Analysis DNA Chromatin High-Throughput Screening Assays Histone Code Histone biology.protein Chromatin Immunoprecipitation Sequencing Protein Processing Post-Translational Chromatin immunoprecipitation 030217 neurology & neurosurgery |
| Zdroj: | Nat Protoc |
| ISSN: | 1750-2799 1754-2189 |
| Popis: | Epigenetic mechanisms such as histone modifications play critical roles in adaptive tuning of chromatin structures. Profiling of various histone modifications at the genome scale using 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 cell type, cell isolation from tissues is often necessary to generate a homogeneous cell population, and such operations tend to yield a low number of cells. In addition, high-throughput processing is often desirable because of the multiplexity of histone marks of interest and the large quantity of samples in a hospital setting. In this protocol, we provide detailed instructions for device fabrication, setup, and operation of microfluidic oscillatory washing-based chromatin immunoprecipitation followed by sequencing (MOWChIP-seq) for profiling of histone modifications using as few as 100 cells per assay with a throughput as high as eight assays in one run. MOWChIP-seq operation involves flowing of chromatin fragments through a packed bed of antibody-coated beads, followed by vigorous microfluidic oscillatory washing. Our process is semi-automated to reduce labor and improve reproducibility. Using one eight-unit device, it takes 2 d to produce eight sequencing libraries from chromatin samples. The technology is scalable. We used the protocol to study a number of histone modifications in various types of mouse and human tissues. The protocol can be conducted by a user who is familiar with molecular biology procedures and has basic engineering skills. |
| Databáze: | OpenAIRE |
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