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DNA Damage In Situ Ligation Followed by Proximity Ligation Assay (DI-PLA)

  • Alessandro Galbiati
  • Fabrizio d’Adda di Fagagna
Part of the Methods in Molecular Biology book series (MIMB, volume 1896)

Abstract

Cells have evolved DNA repair mechanisms to maintain their genetic information unaltered and a DNA damage response pathway that coordinates DNA repair with several cellular events. Despite a clear role for DNA damage in the form of DNA double-strand breaks (DSBs) in several cellular processes, the most commonly used methods to detect DNA lesions are indirect, and rely on antibody-based recognition of DNA damage-associated factors, leaving several important questions unanswered. Differently, here we describe DNA damage In situ ligation followed by Proximity Ligation Assay (DI-PLA), that allows sensitive detection of physical DSBs in fixed cells, through direct labeling of the DSBs with biotinylated oligonucleotides, and subsequent signal amplification by PLA between biotin and a partner protein in the proximity of the DNA break.

Key words

DI-PLA PLA Single-cell Imaging DNA damage response (DDR) DNA damage DNA double-strand break (DSB) 

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Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2019

Authors and Affiliations

  1. 1.Oncology IMEDAstraZeneca UK LtdCambridgeUK
  2. 2.IFOM-FoundationThe FIRC Institute of Molecular Oncology FoundationMilanItaly
  3. 3.Istituto di Genetica Molecolare, Consiglio Nazionale delle RicerchePaviaItaly

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