title: Standardizing chromatin research: a simple and universal method for ChIP-seq.
abstract: Chromatin immunoprecipitation followed by next generation sequencing (ChIP-seq) is a key technique in chromatin research. Although heavily applied, existing ChIP-seq protocols are often highly fine-tuned workflows, optimized for specific experimental requirements. Especially the initial steps of ChIP-seq, particularly chromatin shearing, are deemed to be exceedingly cell-type-specific, thus impeding any protocol standardization efforts. Here we demonstrate that harmonization of ChIP-seq workflows across cell types and conditions is possible when obtaining chromatin from properly isolated nuclei. We established an ultrasound-based nuclei extraction method (NEXSON: Nuclei EXtraction by SONication) that is highly effective across various organisms, cell types and cell numbers. The described method has the potential to replace complex cell-type-specific, but largely ineffective, nuclei isolation protocols. By including NEXSON in ChIP-seq workflows, we completely eliminate the need for extensive optimization and sample-dependent adjustments. Apart from this significant simplification, our approach also provides the basis for a fully standardized ChIP-seq and yields highly reproducible transcription factor and histone modifications maps for a wide range of different cell types. Even small cell numbers (∼10,000 cells per ChIP) can be easily processed without application of modified chromatin or library preparation protocols.
authors.affiliation: Max Planck Institute of Immunobiology and Epigenetics, Stübeweg 51, Freiburg, 79108, Germany arrigoni@ie-freiburg.mpg.de.
authors.forename: Laura
authors.identifier: http://orcid.org/0000-0002-3626-4468
authors.initials: L
authors.lastname: Arrigoni
authors.affiliation: Max Planck Institute of Immunobiology and Epigenetics, Stübeweg 51, Freiburg, 79108, Germany.
authors.forename: Andreas S
authors.identifier: http://orcid.org/0000-0002-4166-0991
authors.initials: AS
authors.lastname: Richter
authors.affiliation: Max Planck Institute of Immunobiology and Epigenetics, Stübeweg 51, Freiburg, 79108, Germany.
authors.forename: Emily
authors.initials: E
authors.lastname: Betancourt
authors.affiliation: Max Planck Institute of Immunobiology and Epigenetics, Stübeweg 51, Freiburg, 79108, Germany.
authors.forename: Kerstin
authors.initials: K
authors.lastname: Bruder
authors.affiliation: Luxembourg Centre for Systems Biomedicine, Université du Luxembourg, avenue du Swing 6, Belvaux, 4366, Luxembourg.
authors.forename: Sarah
authors.initials: S
authors.lastname: Diehl
authors.affiliation: Max Planck Institute of Immunobiology and Epigenetics, Stübeweg 51, Freiburg, 79108, Germany.
authors.forename: Thomas
authors.initials: T
authors.lastname: Manke
authors.affiliation: Max Planck Institute of Immunobiology and Epigenetics, Stübeweg 51, Freiburg, 79108, Germany boenisch@ie-freiburg.mpg.de.
authors.forename: Ulrike
authors.initials: U
authors.lastname: Bönisch
mesh_terms: D000818:Animals; D002458:Cell Fractionation / Q000379:methods / Q000592:standards*; D002460:Cell Line; D002467:Cell Nucleus / Q000235:genetics; D002478:Cells, Cultured; D002843:Chromatin / Q000302:isolation & purification; D047369:Chromatin Immunoprecipitation / Q000379:methods*; D005260:Female; D056945:Hep G2 Cells; D059014:High-Throughput Nucleotide Sequencing / Q000379:methods*; D006657:Histones / Q000378:metabolism; D006801:Humans; D008297:Male; D051379:Mice; D015203:Reproducibility of Results; D017422:Sequence Analysis, DNA / Q000379:methods*; D013010:Sonication; D014157:Transcription Factors / Q000378:metabolism
chemical_list: D002843:Chromatin; D006657:Histones; D014157:Transcription Factors