Genome-wide and sister chromatid-resolved profiling of protein occupancy in replicated chromatin with ChOR-seq and SCAR-seq
| Autor: | Petryk, Nataliya, Reverón Gómez, Nazaret, González-Aguilera, Cristina, Dalby, Maria, Andersson, Robin, Groth, Anja |
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| Přispěvatelé: | Independent Research Fund Denmark, European Research Council, Lundbeck Foundation, Novo Nordisk Foundation, Université de Paris, Agence Nationale de la Recherche (France), Ministerio de Ciencia, Innovación y Universidades (España), Agencia Estatal de Investigación (España), Universidad de Sevilla |
| Rok vydání: | 2021 |
| Zdroj: | Digital.CSIC. Repositorio Institucional del CSIC instname |
| Popis: | Elucidating the mechanisms underlying chromatin maintenance upon genome replication is critical for the understanding of how gene expression programs and cell identity are preserved across cell divisions. Here, we describe two recently developed techniques, chromatin occupancy after replication (ChOR)-seq and sister chromatids after replication (SCAR)-seq, that profile chromatin occupancy on newly replicated DNA in mammalian cells in 5 d of bench work. Both techniques share a common strategy that includes pulse labeling of newly synthesized DNA and chromatin immunoprecipitation (ChIP), followed by purification and high-throughput sequencing. Whereas ChOR-seq quantitatively profiles the post-replicative abundance of histone modifications and chromatin-associated proteins, SCAR-seq distinguishes chromatin occupancy between nascent sister chromatids. Together, these two complementary techniques have unraveled key mechanisms controlling the inheritance of modified histones during replication and revealed locus-specific dynamics of histone modifications across the cell cycle. Here, we provide the experimental protocols and bioinformatic pipelines for these methods. Research in the Groth laboratory was supported by the Independent Research Fund Denmark (7016-00042B and 4092-00404), the European Research Council (CoG no. 724436) and the Lundbeck Foundation (R198-2015-269 and R165-2013-15306). Research at CPR is supported by the Novo Nordisk Foundation (NNF14CC0001). N.P. is supported by LabEx ‘Who Am I?’ #ANR-11-LABX-0071 and the Université de Paris IdEx #ANR-18-IDEX-0001 funded by the French Government through its ‘Investments for the Future’ program and by an INTEGER grant from Agence Nationale de la Recherche (ANR-19-CE12-0030-01). C.G.-A. is the recipient of a Ramón y Cajal Contract RYC2018-025485-I, and her laboratory is supported by grants PID2019-105742GA-100 from the Spanish Government and VIPPIT-2019-IV.2 from the University of Sevilla. M.D. and R.A. were supported by the Independent Research Fund Denmark (6108-00038B) and the European Research Council (StG no. 638173) awarded to R.A. |
| Databáze: | OpenAIRE |
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