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Non-saturated nucleic acid probes with a broad dynamic range.

Xinmiao Kang1, Yu Liu1, Dandan Tian1

  • 1State Key Laboratory of Organic-Inorganic Composites, Beijing Advanced Innovation Center for Soft Matter Science and Engineering, Beijing Key Laboratory of Bioprocess, College of Life Science and Technology Beijing University of Chemical Technology, Beijing, 100029, China. xinsu@mail.buct.edu.cn.

Nanoscale Horizons
|June 23, 2025
PubMed
Summary
This summary is machine-generated.

Researchers developed a novel non-saturating nucleic acid probe (NSNAP) that overcomes limitations of traditional molecular probes. This reusable probe significantly expands detection dynamic range and enables sensitive quantification of viral and bacterial genes.

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Area of Science:

  • Molecular biology
  • Biotechnology
  • Analytical chemistry

Background:

  • Traditional molecular probes have limited dynamic range and reusability due to irreversible target binding.
  • Signal saturation occurs when target concentration exceeds probe concentration, hindering accurate quantification.
  • Existing probes require higher concentrations than targets, limiting sensitivity and increasing costs.

Purpose of the Study:

  • To develop a novel non-saturating nucleic acid probe (NSNAP) overcoming limitations of conventional probes.
  • To enhance the dynamic range, reusability, and sensitivity of molecular detection systems.
  • To demonstrate the efficacy of NSNAP in quantifying clinically relevant genes in complex biological samples.

Main Methods:

  • Designed NSNAP integrating an affinity probe with a target-degrading enzyme for continuous signaling.
  • Utilized enzymes like Exonuclease III or λ Exonucleases for probe resetting and target degradation.
  • Validated NSNAP performance by quantifying viral (HIV, HHV, HPV) and bacterial (oprL, dnaJ, ddl) genes from 1 to 1000 fM.

Main Results:

  • Achieved a 5000-fold increase in dynamic range compared to conventional probes (81-fold).
  • Enabled detection of targets at concentrations 250 times greater than probe concentrations.
  • Demonstrated at least seven cycles of NSNAP reuse with consistent performance.
  • Successfully quantified target genes in complex biological matrices with strong linear correlations.

Conclusions:

  • NSNAP technology offers a significant advancement over traditional molecular probes.
  • The non-saturating and reusable nature of NSNAP enhances cost-effectiveness and sustainability.
  • NSNAP holds great potential for developing advanced DNA nanotechnology-based diagnostic tools for research and clinical applications.