A versatile system to introduce clusters of genomic double-strand breaks in large cell populations.
DNA damage
chromothripsis
genomic rearrangements
Journal
Genes, chromosomes & cancer
ISSN: 1098-2264
Titre abrégé: Genes Chromosomes Cancer
Pays: United States
ID NLM: 9007329
Informations de publication
Date de publication:
05 2021
05 2021
Historique:
revised:
19
07
2020
received:
10
05
2020
accepted:
21
07
2020
pubmed:
1
8
2020
medline:
16
2
2022
entrez:
1
8
2020
Statut:
ppublish
Résumé
In vitro assays for clustered DNA lesions will facilitate the analysis of the mechanisms underlying complex genome rearrangements such as chromothripsis, including the recruitment of repair factors to sites of DNA double-strand breaks (DSBs). We present a novel method generating localized DNA DSBs using UV irradiation with photomasks. The size of the damage foci and the spacing between lesions are fully adjustable, making the assay suitable for different cell types and targeted areas. We validated this setup with genomically stable epithelial cells, normal fibroblasts, pluripotent stem cells, and patient-derived primary cultures. Our method does not require a specialized device such as a laser, making it accessible to a broad range of users. Sensitization by 5-bromo-2-deoxyuridine incorporation is not required, which enables analyzing the DNA damage response in post-mitotic cells. Irradiated cells can be cultivated further, followed by time-lapse imaging or used for downstream biochemical analyses, thanks to the high throughput of the system. Importantly, we showed genome rearrangements in the irradiated cells, providing a proof of principle for the induction of structural variants by localized DNA lesions.
Types de publication
Journal Article
Research Support, Non-U.S. Gov't
Langues
eng
Sous-ensembles de citation
IM
Pagination
303-313Informations de copyright
© 2020 The Authors. Genes, Chromosomes and Cancer published by Wiley Periodicals LLC.
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