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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.
Denatured DNA fragments must be transferred onto a carrier membrane from the gel to make it accessible to a probe - a small ssDNA fragment complementary to the target DNA...
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...

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Selective nucleic acid capture with shielded covalent probes.

Jeffrey R Vieregg1, Hosea M Nelson, Brian M Stoltz

  • 1Department of Bioengineering, California Institute of Technology, Pasadena, California 91125, USA.

Journal of the American Chemical Society
|June 11, 2013
PubMed
Summary
This summary is machine-generated.

Shielded covalent (SC) probes offer enhanced selectivity and durable nucleic acid capture by combining base pairing with covalent cross-linking. These probes overcome limitations of traditional methods, enabling stable binding even under disruptive conditions.

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Area of Science:

  • Molecular Biology
  • Biochemistry
  • Nucleic Acid Chemistry

Background:

  • Traditional nucleic acid probes rely on base pairing, leading to limitations in selectivity and affinity.
  • Imperfect selectivity and reversible binding hinder the efficiency and stability of conventional probes.
  • A fundamental trade-off exists between probe selectivity and the durability of target molecule capture.

Purpose of the Study:

  • To introduce and characterize Shielded Covalent (SC) probes that overcome limitations of traditional nucleic acid probes.
  • To demonstrate the ability of SC probes to achieve both high selectivity and durable target capture.
  • To explore the potential applications of SC probes in various biological settings.

Main Methods:

  • Development of SC probes utilizing programmable base pairing and molecular conformation changes for selectivity.
  • Incorporation of activatable covalent cross-linking for durable target capture.
  • Testing probe performance with DNA/RNA oligonucleotides and full-length mRNA targets in various conditions.

Main Results:

  • SC probes achieved near-quantitative covalent capture of complementary DNA/RNA targets with high discrimination against mismatched targets (2-3 orders of magnitude).
  • Selective covalent capture was demonstrated for full-length mRNA targets, comparable to oligonucleotide capture.
  • Single-nucleotide mismatches were efficiently rejected (1-2 orders of magnitude discrimination), and cross-links were reversible.

Conclusions:

  • SC probes successfully combine high sequence selectivity with durable target retention, even under denaturing conditions.
  • This novel probe chemistry overcomes the traditional trade-off between selectivity and stable binding.
  • SC probes offer diverse applications for selective and stable nucleic acid detection in challenging environments.