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Updated: Jul 5, 2026
12:07
Sequence-specific Labeling of Nucleic Acids and Proteins with Methyltransferases and Cofactor Analogues
Published on: November 22, 2014
Enzymatic labeling of DNA.
Current Protocols in Human Genetics
|April 23, 2008
Summary
This study details methods for creating radioactive probes using nick translation and random priming. It also covers oligonucleotide end-labeling and purification techniques for accurate radioactive labeling.
Area of Science:
- Molecular Biology
- Biochemistry
- Nucleic Acid Chemistry
Background:
- Radioactive probes are essential tools in molecular biology for detecting specific nucleic acid sequences.
- Accurate preparation and characterization of these probes are critical for reliable experimental results.
Purpose of the Study:
- To provide detailed protocols for the preparation of uniformly labeled radioactive probes.
- To describe methods for end-labeling oligonucleotides.
- To outline techniques for purifying probes and measuring their specific activity.
Main Methods:
- Uniformly labeled radioactive probes were prepared using nick translation and random oligonucleotide-primed synthesis.
- Oligonucleotides were end-labeled using T4 polynucleotide kinase.
- Unincorporated deoxynucleotide triphosphate (dNTP) precursors were removed using spin columns.
- Probe specific activity was measured by acid precipitation.
Main Results:
- Established protocols for efficient synthesis of uniformly labeled radioactive probes.
- Successfully demonstrated end-labeling of oligonucleotides.
- Validated methods for purification of labeled probes.
- Quantified probe specific activity using acid precipitation.
Conclusions:
- The described methods provide a comprehensive approach to radioactive probe preparation and characterization.
- These protocols are valuable for researchers requiring high-quality labeled nucleic acid probes.
- Accurate probe preparation and measurement are crucial for applications in molecular biology and diagnostics.
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