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Related Concept Videos

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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In situ hybridization (ISH) is a technique used to detect and localize specific DNA or RNA molecules in cells, tissue, or tissue sections using a labeled probe. The technique was first used in 1969 for the investigation of nucleic acids. It is currently an essential tool in scientific research and clinical settings, especially for diagnostic purposes.
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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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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.
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Visualization of Surface-tethered Large DNA Molecules with a Fluorescent Protein DNA Binding Peptide
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Comparison of methods for generating planar DNA-modified surfaces for hybridization studies.

Amal Kasry1, Paola Borri, Philip R Davies

  • 1Cardiff School of Biosciences, Biomedical Science Building, Museum Avenue, Cardiff CF10 3AX, UK. kasrya@cf.ac.uk

ACS Applied Materials & Interfaces
|April 2, 2010
PubMed
Summary

Researchers compared DNA immobilization methods for sensor technologies. The biotin-streptavidin method offers superior oligonucleotide accessibility and density on glass surfaces compared to direct coupling.

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Area of Science:

  • Biotechnology
  • Surface Chemistry
  • Sensor Technology

Background:

  • Oligonucleotide surface conformation and accessibility are critical for nucleic acid sensor performance.
  • Effective immobilization strategies are needed to maximize the utility of immobilized DNA in biosensors.

Purpose of the Study:

  • To compare two DNA immobilization techniques on glass surfaces.
  • To identify a method yielding high surface density and hybridization accessibility of oligonucleotides in a planar monolayer.

Main Methods:

  • Utilized a combination of analytical techniques, including atomic force microscopy (AFM) and X-ray photoelectron spectroscopy (XPS).
  • Compared direct silanized DNA coupling versus biotin-streptavidin-mediated immobilization.
  • Assessed surface accessibility through hybridization measurements.

Main Results:

  • Direct coupling resulted in high immobilization density (0.013 molecules/nm2) but low accessibility (<15%).
  • Biotin-streptavidin interaction yielded high density (0.02 molecules/nm2) and high accessibility (90%).
  • Both methods demonstrated uniform surface characteristics via AFM and XPS.

Conclusions:

  • The biotin-streptavidin method significantly enhances oligonucleotide accessibility for hybridization on sensor surfaces.
  • This method allows for controlled intermolecular distances between hybridized molecules.
  • The findings are crucial for advancing nucleic acid-based sensor technologies.