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

  • Alessandro GalbiatiEmail author
  • Fabrizio d’Adda di FagagnaEmail author
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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