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Related Concept Videos

Labeling DNA Probes03:31

Labeling DNA Probes

DNA probes are fragments of DNA labeled with a reporter tag to enable their detection or purification. The resulting labeled DNA probes can then hybridize to target nucleic acid sequences through complementary base-pairing, and may be used to recover or identify these regions.
Radioisotopes, fluorophores, or small molecule binding partners like biotin or digoxigenin, are the most widely used reporter tags for labeling DNA probes. These labels can be attached to the probe DNA molecule via...
Single-Strand DNA Binding Proteins01:03

Single-Strand DNA Binding Proteins

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...
Southern Blot02:57

Southern Blot

Agarose gel electrophoresis is very useful in separating DNA fragments by size. Running a DNA ladder containing fragments of the known length alongside the sample helps determine the approximate length of the sample DNA fragments. However, additional steps are needed to verify the sequence identity of the sample DNA fragments.
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FISH - Fluorescent In-situ Hybridization02:07

FISH - Fluorescent In-situ Hybridization

Fluorescence in situ hybridization, or FISH, was developed in the early 1980s and has quickly become one of the most widely used techniques in cytogenetics. Labeled probes are used to bind complementary DNA or RNA sequences on a chromosome or in a region within a cell. Earlier, the probes could only be obtained by cloning or reverse transcription of a DNA template. Currently, the probe oligonucleotides can be synthesized synthetically. Additionally, with the advancement of optical techniques,...

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Related Experiment Video

Updated: May 18, 2026

Split Hybridization Probe Utilizing a DNA Fluorescent Light-up Aptamer as a Signal Reporter for Sequence-Specific Nucleic Acid Analysis
07:10

Split Hybridization Probe Utilizing a DNA Fluorescent Light-up Aptamer as a Signal Reporter for Sequence-Specific Nucleic Acid Analysis

Published on: July 8, 2025

Fluorescent SSB as a reagentless biosensor for single-stranded DNA.

Katy Hedgethorne1, Martin R Webb

  • 1MRC National Institute for Medical Research, London, UK.

Methods in Molecular Biology (Clifton, N.J.)
|September 15, 2012
PubMed
Summary
This summary is machine-generated.

This study introduces a novel reagentless biosensor using a modified single-stranded DNA-binding protein (SSB) to monitor helicase activity. The assay provides a sensitive method for studying DNA unwinding enzymes at different scales.

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A Microfluidic-based Electrochemical Biochip for Label-free DNA Hybridization Analysis
14:53

A Microfluidic-based Electrochemical Biochip for Label-free DNA Hybridization Analysis

Published on: September 10, 2014

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Last Updated: May 18, 2026

Split Hybridization Probe Utilizing a DNA Fluorescent Light-up Aptamer as a Signal Reporter for Sequence-Specific Nucleic Acid Analysis
07:10

Split Hybridization Probe Utilizing a DNA Fluorescent Light-up Aptamer as a Signal Reporter for Sequence-Specific Nucleic Acid Analysis

Published on: July 8, 2025

A Microfluidic-based Electrochemical Biochip for Label-free DNA Hybridization Analysis
14:53

A Microfluidic-based Electrochemical Biochip for Label-free DNA Hybridization Analysis

Published on: September 10, 2014

Area of Science:

  • Biochemistry
  • Molecular Biology
  • Enzymology

Background:

  • Helicases are crucial enzymes that unwind nucleic acid duplexes by hydrolyzing ATP.
  • Studying helicase activity requires diverse biochemical assays to track reaction progress.
  • Existing methods may require specific reagents or lack sensitivity for certain applications.

Purpose of the Study:

  • To develop a novel, reagentless biosensor for assaying helicase activity.
  • To utilize the specific binding properties of single-stranded DNA-binding protein (SSB) for signal generation.
  • To enable the study of DNA unwinding at both bulk and single-molecule levels.

Main Methods:

  • Engineering a fluorescently labeled single-stranded DNA-binding protein (SSB) from Escherichia coli.
  • Utilizing the fluorescence enhancement of the labeled SSB upon binding to single-stranded DNA.
  • Developing a protocol for monitoring helicase-driven DNA unwinding in real-time.
  • Adapting the assay for single-molecule measurements of DNA unwinding.

Main Results:

  • The diethylaminocoumarin-labeled (G26C)SSB exhibited a ~5-fold fluorescence increase upon binding to single-stranded DNA.
  • The developed biosensor effectively monitored the progress of helicase activity on double-stranded DNA.
  • A variant of the assay was successfully applied to single-molecule studies of DNA unwinding.

Conclusions:

  • The fluorescently labeled SSB serves as an effective reagentless biosensor for helicase activity.
  • This assay offers a sensitive and versatile tool for biochemical and biophysical studies of DNA unwinding.
  • The method facilitates the investigation of helicase mechanisms at various scales, from bulk reactions to single molecules.