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Related Concept Videos

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...
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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...
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For successful DNA replication, the unwinding of double-stranded DNA must be accompanied by stabilization and protection of the separated single strands of the DNA. This crucial task is performed by single-strand DNA-binding (SSB) proteins. They bind to the DNA in a sequence-independent manner, which means that the nitrogenous bases of the DNA need not be present in a specific order for binding of SSB proteins to it. The binding of SSB proteins straightens single-stranded DNA (ssDNA) and makes...
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In response to DNA damage, cells can pause the cell cycle to assess and repair the breaks. However, the cell must check the DNA at certain critical stages during the cell cycle. If the cell cycle pauses before DNA replication, the cells will contain twice the amount of DNA. On the other hand, if cells arrest after DNA replication but before mitosis, they will contain four times the normal amount of DNA. With a host of specialized proteins at their disposal,cells must use the right protein at...
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In response to DNA damage, cells can pause the cell cycle to assess and repair the breaks. However, the cell must check the DNA at certain critical stages during the cell cycle. If the cell cycle pauses before DNA replication, the cells will contain twice the amount of DNA. On the other hand, if cells arrest after DNA replication but before mitosis, they will contain four times the normal amount of DNA. With a host of specialized proteins at their disposal,cells must use the right protein at...
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Two- and Three-Dimensional Live Cell Imaging of DNA Damage Response Proteins
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Two- and Three-Dimensional Live Cell Imaging of DNA Damage Response Proteins

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Proteins pinpoint double strand breaks.

Michael M Cox1

  • 1is at the Department of Biochemistry , University of Wisconsin-Madison , Madison , United States cox@biochem.wisc.edu.

Elife
|October 31, 2013
PubMed
Summary
This summary is machine-generated.

Scientists developed a new method to detect DNA double-strand breaks in living bacteria. This technique uses green fluorescent protein and a specialized DNA-binding protein for high-efficiency damage detection.

Keywords:
DNA double-strand breaksE. coliGFPHumanMouseendogenous DNA damagefluorescent-protein fusionsspontaneous DNA breakssynthetic biology

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

  • Microbiology
  • Molecular Biology
  • Genetics

Background:

  • DNA double-strand breaks are a critical form of DNA damage.
  • Detecting these breaks in real-time within living organisms is challenging.
  • Existing methods may lack efficiency or applicability to live-cell imaging.

Purpose of the Study:

  • To develop a highly efficient method for detecting DNA double-strand breaks in living bacteria.
  • To visualize and quantify DNA damage in situ using a novel biosensor.

Main Methods:

  • Constructed a fusion protein combining green fluorescent protein (GFP) with a DNA double-strand break-binding protein.
  • Introduced the biosensor into live bacterial cells.
  • Utilized fluorescence microscopy to observe and quantify GFP signal localization, indicating DNA break sites.

Main Results:

  • The developed biosensor efficiently detected DNA double-strand breaks in living bacteria.
  • High-efficiency localization of the fluorescent signal was observed at sites of DNA damage.
  • The method proved effective for real-time monitoring of DNA damage.

Conclusions:

  • This novel GFP-based biosensor provides a powerful tool for studying DNA double-strand breaks in live bacteria.
  • The technique offers high sensitivity and efficiency for DNA damage detection.
  • Facilitates research into DNA repair mechanisms and genotoxicity testing in bacterial models.