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Related Experiment Video

Updated: Jun 21, 2025

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One-base-mismatch CRISPR-based transistors for single nucleotide resolution assay.

Hongwenjie Ma1, Yicheng Tian2, Derong Kong1

  • 1State Key Laboratory of Molecular Engineering of Polymers, Department of Macromolecular Science, Fudan University, Shanghai, 200433, PR China.

Biosensors & Bioelectronics
|July 10, 2024
PubMed
Summary

This study introduces a CRISPR-based transistor biosensor for rapid and selective single nucleotide variant (SNV) detection in viral RNA. The novel device achieves high sensitivity without amplification, enabling quick diagnostics for respiratory viruses.

Keywords:
BiosensorCRISPR-Cas13aEpidemic detectionGraphene field-effect transistorSingle nucleotide variants

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Area of Science:

  • Biotechnology
  • Molecular Diagnostics
  • Biosensor Technology

Background:

  • Accurate single nucleotide variant (SNV) detection is crucial for genetic research and diagnostics.
  • Current SNV detection methods often require stringent amplification conditions and complex instrumentation.
  • Achieving a balance between sensitivity and selectivity in SNV discrimination remains a challenge.

Purpose of the Study:

  • To develop a rapid and highly selective biosensor for detecting SNVs in viral RNA.
  • To engineer a CRISPR-Cas13a system for direct capture of target SNV RNA on a transistor surface.
  • To demonstrate the utility of this biosensor for clinical diagnostics of respiratory viruses.

Main Methods:

  • A CRISPR-based transistor biosensor utilizing CRISPR-Cas13a engineered with a synthetic mismatch in crRNA.
  • Direct capture of target SNV RNA on a graphene channel surface, inducing an electrical signal.
  • Detection without the need for nucleic acid amplification or reporter molecules.

Main Results:

  • The biosensor achieved a detection limit as low as 5 copies/100 μL, comparable to real-time quantitative PCR.
  • Operational range spans 10 to 5 × 10^5 copies/mL in artificial saliva.
  • Discrimination between wild-type and SNV RNA was achieved within 15 minutes.
  • Successfully detected respiratory viruses, including influenza A, B, and SARS-CoV-2 variants, in 19 oropharyngeal specimens.

Conclusions:

  • The CRISPR-transistor biosensor offers a highly accurate and sensitive method for SNV detection.
  • This technology obviates the need for amplification and reporter molecules, simplifying the detection process.
  • The biosensor is suitable for field-deployable nucleic acid screening and diagnostics of viral infections.