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Related Concept Videos

Homologous Recombination02:31

Homologous Recombination

The basic reaction of homologous recombination (HR) involves two chromatids that contain DNA sequences sharing a significant stretch of identity. One of these sequences uses a strand from another as a template to synthesize DNA in an enzyme-catalyzed reaction. The final product is a novel amalgamation of the two substrates. To ensure an accurate recombination of sequences, HR is restricted to the S and G2 phases of the cell cycle. At these stages, the DNA has been replicated already and the...
Fixing Double-strand Breaks02:04

Fixing Double-strand Breaks

The double-stranded structure of DNA has two major advantages. First, it serves as a safe repository of genetic information where one strand serves as the back-up in case the other strand is damaged. Second, the double-helical structure can be wrapped around proteins called histones to form nucleosomes, which can then be tightly wound to form chromosomes. This way, DNA chains up to 2 inches long can be contained within microscopic structures in a cell. A double-stranded break not only damages...
Fixing Double-strand Breaks02:04

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The double-stranded structure of DNA has two major advantages. First, it serves as a safe repository of genetic information where one strand serves as the back-up in case the other strand is damaged. Second, the double-helical structure can be wrapped around proteins called histones to form nucleosomes, which can then be tightly wound to form chromosomes. This way, DNA chains up to 2 inches long can be contained within microscopic structures in a cell. A double-stranded break not only damages...
Nucleotide Excision Repair01:08

Nucleotide Excision Repair

Overview
Nucleotide Excision Repair01:38

Nucleotide Excision Repair

DNA Distortion and Damage
Cells are regularly exposed to mutagens—factors in the environment that can damage DNA and generate mutations. UV radiation is one of the most common mutagens and is estimated to introduce a significant number of changes in DNA. These include bends or kinks in the structure, which can block DNA replication or transcription. If these errors are not fixed, the damage can cause mutations, which in turn can result in cancer or disease depending on which sequences are...
Long-patch Base Excision Repair01:02

Long-patch Base Excision Repair

Since the discovery of the two BER pathways, there has been a debate about how a cell chooses one pathway over the other and the factors determining this selection. Numerous in vitro experiments have pointed out multiple determinants for the sub-pathway selection. These are:

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Related Experiment Video

Updated: Jun 4, 2026

Application of Laser Micro-irradiation for Examination of Single and Double Strand Break Repair in Mammalian Cells
08:18

Application of Laser Micro-irradiation for Examination of Single and Double Strand Break Repair in Mammalian Cells

Published on: September 5, 2017

Creating localized DNA double-strand breaks with microirradiation.

Keiji Suzuki1, Motohiro Yamauchi, Yasuyoshi Oka

  • 1Atomic Bomb Disease Institute, Course of Life Sciences and Radiation Research, Nagasaki University Graduate School of Biomedical Sciences, Nagasaki, Japan. kzsuzuki@nagasaki-u.ac.jp

Nature Protocols
|February 5, 2011
PubMed
Summary

This study presents a simple method for creating localized DNA double-strand breaks (DSBs) using 5-bromo-2'-deoxyuridine (BrdU) labeling and ultraviolet C (UVC) irradiation. This technique allows for easy visualization of DNA repair and damage response proteins.

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Application of Laser Micro-irradiation for Examination of Single and Double Strand Break Repair in Mammalian Cells
08:18

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Characterizing DNA Repair Processes at Transient and Long-lasting Double-strand DNA Breaks by Immunofluorescence Microscopy
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Characterizing DNA Repair Processes at Transient and Long-lasting Double-strand DNA Breaks by Immunofluorescence Microscopy

Published on: June 8, 2018

Area of Science:

  • Molecular Biology
  • Cell Biology
  • Biochemistry

Background:

  • Localized DNA double-strand breaks (DSBs) are crucial for studying DNA repair and damage response pathways.
  • Existing methods for inducing DSBs, such as laser microirradiation, often require specialized and expensive equipment.

Purpose of the Study:

  • To develop a cost-effective and accessible protocol for generating localized DSBs in cells.
  • To enable visualization of protein recruitment involved in DNA repair and damage response without specialized equipment.

Main Methods:

  • The protocol involves labeling cells with 5-bromo-2 -deoxyuridine (BrdU) for 48-72 hours.
  • Cells are then exposed to ultraviolet C (UVC) irradiation through microporous polycarbonate membranes.
  • This induces localized DSBs within subnuclear regions, independent of the cell cycle phase.

Main Results:

  • The BrdU-labeling and UVC irradiation method effectively creates localized DSBs.
  • Recruitment of DNA repair, DNA damage response, chromatin remodeling, and histone modification proteins can be visualized.
  • DSBs become detectable within 30 minutes post-irradiation.
  • The quality of DSBs generated is comparable to that achieved with laser microirradiation.

Conclusions:

  • This protocol provides a simple, minimal-requirement method for generating localized DSBs.
  • It facilitates the study of DNA damage response and repair mechanisms in various cell biology and molecular biology applications.
  • The technique allows for the visualization of key protein dynamics at DSB sites without specialized equipment.