Genome-wide mapping of DNA double-strand breaks from eukaryotic cell cultures using Break-seq

Genome-wide mapping of DNA double-strand breaks from eukaryotic cell cultures using Break-seq

Summary: We describe a genome-wide DNA double-strand break (DSB) mapping technique, Break-seq. In this protocol, we provide step-by-step instructions for cell embedment in agarose, in-gel DSB labeling and subsequent capture, followed by standard Illumina library construction and sequencing. We also...

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Bibliographic Details
Main Authors: Ishita Joshi, Jenna DeRycke, Megan Palmowski, Robert LeSuer, Wenyi Feng
Format: Article
Language:English
Published: Elsevier 2021-06-01
Series:STAR Protocols
Subjects:
Genomics
Sequencing
Model Organisms
Molecular Biology
Online Access:http://www.sciencedirect.com/science/article/pii/S2666166721002616
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spelling doaj-12d0c7d64e33488eb0851ac7e4883c1e2021-06-21T04:25:40ZengElsevierSTAR Protocols2666-16672021-06-0122100554Genome-wide mapping of DNA double-strand breaks from eukaryotic cell cultures using Break-seqIshita Joshi0Jenna DeRycke1Megan Palmowski2Robert LeSuer3Wenyi Feng4Department of Biochemistry and Molecular Biology, SUNY Upstate Medical University, Syracuse, NY 13210, USA; Corresponding authorDepartment of Chemistry and Biochemistry, SUNY Brockport, Brockport, NY 14420, USADepartment of Chemistry and Biochemistry, SUNY Brockport, Brockport, NY 14420, USADepartment of Chemistry and Biochemistry, SUNY Brockport, Brockport, NY 14420, USADepartment of Biochemistry and Molecular Biology, SUNY Upstate Medical University, Syracuse, NY 13210, USA; Corresponding authorSummary: We describe a genome-wide DNA double-strand break (DSB) mapping technique, Break-seq. In this protocol, we provide step-by-step instructions for cell embedment in agarose, in-gel DSB labeling and subsequent capture, followed by standard Illumina library construction and sequencing. We also provide the framework for sequence data processing and DSB peak identification. Finally, we present a custom-designed 3D-printed device for processing agarose-embedded DNA samples. The protocol is applicable to Saccharomyces cerevisiae, as well as mammalian suspension, adherent, and 3D organoid cell cultures.For complete details on the use and execution of this protocol, please refer to Hoffman et al. (2015) and Chakraborty et al. (2020).http://www.sciencedirect.com/science/article/pii/S2666166721002616GenomicsSequencingModel OrganismsMolecular Biology
collection DOAJ
language English
format Article
sources DOAJ
author Ishita Joshi
Jenna DeRycke
Megan Palmowski
Robert LeSuer
Wenyi Feng
spellingShingle Ishita Joshi
Jenna DeRycke
Megan Palmowski
Robert LeSuer
Wenyi Feng
Genome-wide mapping of DNA double-strand breaks from eukaryotic cell cultures using Break-seq
STAR Protocols
Genomics
Sequencing
Model Organisms
Molecular Biology
author_facet Ishita Joshi
Jenna DeRycke
Megan Palmowski
Robert LeSuer
Wenyi Feng
author_sort Ishita Joshi
title Genome-wide mapping of DNA double-strand breaks from eukaryotic cell cultures using Break-seq
title_short Genome-wide mapping of DNA double-strand breaks from eukaryotic cell cultures using Break-seq
title_full Genome-wide mapping of DNA double-strand breaks from eukaryotic cell cultures using Break-seq
title_fullStr Genome-wide mapping of DNA double-strand breaks from eukaryotic cell cultures using Break-seq
title_full_unstemmed Genome-wide mapping of DNA double-strand breaks from eukaryotic cell cultures using Break-seq
title_sort genome-wide mapping of dna double-strand breaks from eukaryotic cell cultures using break-seq
publisher Elsevier
series STAR Protocols
issn 2666-1667
publishDate 2021-06-01
description Summary: We describe a genome-wide DNA double-strand break (DSB) mapping technique, Break-seq. In this protocol, we provide step-by-step instructions for cell embedment in agarose, in-gel DSB labeling and subsequent capture, followed by standard Illumina library construction and sequencing. We also provide the framework for sequence data processing and DSB peak identification. Finally, we present a custom-designed 3D-printed device for processing agarose-embedded DNA samples. The protocol is applicable to Saccharomyces cerevisiae, as well as mammalian suspension, adherent, and 3D organoid cell cultures.For complete details on the use and execution of this protocol, please refer to Hoffman et al. (2015) and Chakraborty et al. (2020).
topic Genomics
Sequencing
Model Organisms
Molecular Biology
url http://www.sciencedirect.com/science/article/pii/S2666166721002616
work_keys_str_mv AT ishitajoshi genomewidemappingofdnadoublestrandbreaksfromeukaryoticcellculturesusingbreakseq
AT jennaderycke genomewidemappingofdnadoublestrandbreaksfromeukaryoticcellculturesusingbreakseq
AT meganpalmowski genomewidemappingofdnadoublestrandbreaksfromeukaryoticcellculturesusingbreakseq
AT robertlesuer genomewidemappingofdnadoublestrandbreaksfromeukaryoticcellculturesusingbreakseq
AT wenyifeng genomewidemappingofdnadoublestrandbreaksfromeukaryoticcellculturesusingbreakseq
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