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DNA isolation protocols can be fast and straightforward or complex and time-consuming depending on the type and quality of DNA required for further processing. For example, plasmid DNA extraction is a bit more complicated than genomic DNA extraction because of the need for an appropriate lysis method to separate plasmid DNA from gDNA during isolation. However, for specific applications, such as long-range DNA sequencing that require a good yield of high- quality DNA samples, we need to follow...
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A Quantitative Assay to Study Protein:DNA Interactions, Discover Transcriptional Regulators of Gene Expression, and Identify Novel Anti-tumor Agents
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A rapid test for protein-DNA interactions.

Casey J Toft1, Holly M Radford1, Alanna E Sorenson1

  • 1Biomedical Sciences and Molecular Biology, College of Medicine and Dentistry, James Cook University, Douglas QLD 4811, Australia.

Nucleic Acids Research
|February 18, 2026
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Summary
This summary is machine-generated.

We developed a rapid, instrument-free assay called R-PNAI-T to quickly detect protein-DNA interactions. This new method is sensitive, robust, and versatile for various biological research applications.

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Area of Science:

  • Molecular Biology
  • Biotechnology
  • Biochemistry

Background:

  • Characterizing protein-DNA interactions is crucial for understanding biological processes.
  • Existing methods for studying these interactions are often time-consuming and labor-intensive.

Purpose of the Study:

  • To introduce a rapid, instrument-free assay for examining protein-DNA interactions.
  • To validate the assay's sensitivity, robustness, and versatility across different biological contexts.

Main Methods:

  • Development of the rapid protein-DNA interaction test (R-PNAI-T) using GFP-tagged proteins and lateral flow assay principles.
  • Validation with bacterial proteins (replication, transcription) and detection of complexes using a dipstick.
  • Testing assay performance in human serum, bacterial lysates, and in the presence of biotin contaminants.

Main Results:

  • The R-PNAI-T assay detects protein-DNA complexes in approximately 15 minutes.
  • Achieved high sensitivity (∼0.7 fmol) with the Escherichia coli Tus protein.
  • Demonstrated capability to discern subtle differences in protein-DNA binding affinity.
  • Confirmed functional interaction between Burkholderia pseudomallei DnaA protein and its target DNA sequence.

Conclusions:

  • The R-PNAI-T is a versatile, cost-effective, and user-friendly tool for studying protein-DNA interactions.
  • The assay is robust, functional in various biological matrices, and tolerant to contaminants.
  • Offers broad applications in biological research, biotechnology, and quality control processes.