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Related Concept Videos

DNA Isolation01:24

DNA Isolation

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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Self-phosphorylating DNAzyme DK1 enables programmable multi-analyte readout via PfAgo.

Tao Hu1, Hongmei Feng2, Xinxin Ke3

  • 1Children's Hospital, Zhejiang University School of Medicine, National Clinical Research Center for Child Health, Hangzhou, 310052, China; School of Basic Medical Sciences & Forensic Medicine, Hangzhou Medical College, Hangzhou, 310000, China; Department of Biological Sciences, Faculty of Science, National University of Singapore, 117543, Singapore.

Biosensors & Bioelectronics
|September 8, 2025
PubMed
Summary

A novel biosensing platform, RASTEN, integrates DNAzyme kinase 1 (DK1) and PfAgo for rapid, visual detection of ATP and protein biomarkers. This cost-effective, instrument-free assay offers versatile point-of-care diagnostics.

Keywords:
Argonaute nucleaseCascade cleavageDeoxyribozyme kinase 1POCSelf-phosphorylation

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Area of Science:

  • Biochemistry and Molecular Biology
  • Biosensing Technology
  • Diagnostic Assays

Background:

  • DNAzymes with kinase activity, like DNAzyme kinase 1 (DK1), show promise but have limited practical applications.
  • DK1's self-phosphorylation upon ATP binding is a key feature not fully exploited for broad diagnostics.

Purpose of the Study:

  • To develop a novel, programmable cascade sensing platform by coupling DK1 and PfAgo.
  • To enable rapid, visual, and versatile detection of non-nucleic acid and nucleic acid targets.

Main Methods:

  • Integration of DK1's autophosphorylation with PfAgo for a sequential reporter probe cleavage system.
  • Development of a point-of-care (POC) diagnostic assay using test strips for ATP and protein biomarker detection.
  • Demonstration of programmability through modifiable DK1 substrate sequences.

Main Results:

  • Successful establishment of the RASTEN (Robust pfAgo-based Strategy for POC Testing Non-nucleic Acid and Nucleic Acid) platform.
  • Highly sensitive and visual detection of ATP in simulated human serum and protein biomarkers using alkaline phosphatase (ALP) activity.
  • Demonstrated advantages over commercial kits: reduced cost, simplified user experience, and instrument-free operation.

Conclusions:

  • The DK1-PfAgo cascade system provides a robust and adaptable biosensing platform.
  • RASTEN offers versatile, cost-effective, and practical multi-analyte diagnostics for point-of-care applications.
  • This technology significantly advances the field of non-nucleic acid and nucleic acid detection.