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Fluorescence in situ hybridization, or FISH, was developed in the early 1980s and has quickly become one of the most widely used techniques in cytogenetics. Labeled probes are used to bind complementary DNA or RNA sequences on a chromosome or in a region within a cell. Earlier, the probes could only be obtained by cloning or reverse transcription of a DNA template. Currently, the probe oligonucleotides can be synthesized synthetically. Additionally, with the advancement of optical techniques,...

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Field-Deployable Candidatus Liberibacter asiaticus Detection Using Recombinase Polymerase Amplification Combined with CRISPR-Cas12a
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CRISPR/Cas9-Based Lateral Flow and Fluorescence Diagnostics.

Mark J Osborn1, Akshay Bhardwaj1, Samuel P Bingea1

  • 1Department of Pediatrics, Division of Blood and Marrow Transplant & Cellular Therapy, University of Minnesota Medical School, Minneapolis, MN 55455, USA.

Bioengineering (Basel, Switzerland)
|March 6, 2021
PubMed
Summary
This summary is machine-generated.

CRISPR/Cas9 technology enables rapid, single-base specific detection of SARS-CoV-2 using a lateral flow assay. A multiplex assay also differentiates between common respiratory viruses, offering scalable point-of-care diagnostics.

Keywords:
CRISPR/Cas9SARS-Co-V2lateral flow assay

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Area of Science:

  • Molecular Biology
  • Biotechnology
  • Infectious Disease Diagnostics

Background:

  • Clustered Regularly Interspaced Short Palindromic Repeat (CRISPR/Cas) systems offer precise nucleic acid targeting capabilities.
  • Accurate and rapid detection of respiratory viral pathogens is crucial for public health, especially for diseases with overlapping symptoms.

Purpose of the Study:

  • To develop a CRISPR/Cas9-based lateral flow assay for sensitive and specific detection of SARS-CoV-2.
  • To create a multiplex fluorescence assay for simultaneous detection and differentiation of multiple respiratory viruses.

Main Methods:

  • Integration of commercially available reagents into a CRISPR/Cas9-based lateral flow assay for SARS-CoV-2 detection.
  • Development of a rapid, multiplex fluorescence CRISPR/Cas9 nuclease cleavage assay for simultaneous detection of SARS-CoV-2, influenza A/B, and RSV.

Main Results:

  • The CRISPR/Cas9 lateral flow assay demonstrated single-base specificity for SARS-CoV-2 detection.
  • The multiplex fluorescence assay successfully detected and differentiated between SARS-CoV-2, influenza A/B, and RSV in a single reaction.
  • Both assays require minimal equipment, facilitating field-based deployment.

Conclusions:

  • CRISPR/Cas9 technology provides a robust platform for point-of-care diagnosis of viral infections.
  • The developed assays offer scalable solutions for identifying respiratory viral pathogens with overlapping clinical presentations.
  • This work demonstrates the potential of CRISPR/Cas9 for rapid, multiplexed diagnostics in resource-limited settings.