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Cas12c-DETECTOR: A specific and sensitive Cas12c-based DNA detection platform.

Zupeng Wang1, Caihong Zhong1

  • 1Key Laboratory of Plant Germplasm Enhancement and Specialty Agriculture, Wuhan Botanical Garden, Chinese Academy of Sciences, Wuhan 430074, China; Innovative Academy of Seed Design, Chinese Academy of Sciences, Beijing 100101, China; Engineering Laboratory for Kiwifruit Industrial Technology, Chinese Academy of Sciences, Wuhan 430074, China.

International Journal of Biological Macromolecules
|October 29, 2021
PubMed
Summary
This summary is machine-generated.

A new CRISPR-based nucleic acid detection platform, Cas12c-DETECTOR, offers robust, sensitive, and precise pathogen identification. This tool recognizes a 5'-TG protospacer adjacent motif (PAM) and aids in diagnosing human and plant diseases.

Keywords:
CRISPRCas12cNucleic acid detectionSNPSensitivity

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Area of Science:

  • Biotechnology
  • Molecular Biology
  • Genetics

Background:

  • Nucleic acid detection platforms are crucial for clinical diagnosis of pathogens and disease control.
  • CRISPR-based tools offer rapid and sensitive detection but face limitations in protospacer adjacent motif (PAM) recognition, specificity, and sensitivity.
  • Existing CRISPR systems often require specific PAM sequences that can limit their broad applicability.

Purpose of the Study:

  • To develop a novel CRISPR-based nucleic acid detection platform with improved characteristics.
  • To overcome the limitations of existing CRISPR detection tools, particularly concerning PAM recognition and specificity.
  • To establish a versatile platform for the sensitive and precise detection of nucleic acids from human and plant pathogens.

Main Methods:

  • Development of a new Cas12c-based nucleic acid detection platform, termed Cas12c-DETECTOR.
  • Characterization of the Cas12c-DETECTOR system, focusing on its protospacer adjacent motif (PAM) recognition (5 acg-TG).
  • Optimization of single-guide RNA (sgRNA) for enhanced specificity and sensitivity.
  • Integration with pre-amplification techniques and detection methods (lateral flow strips, visual fluorescence).

Main Results:

  • Cas12c-DETECTOR successfully recognizes a 5 acg-TG protospacer adjacent motif (PAM).
  • The platform demonstrated high sensitivity and specificity in detecting target nucleic acid sequences.
  • Cas12c-DETECTOR coupled with optimized sgRNA accurately identified single nucleotide polymorphisms (SNPs).
  • The system proved effective for diagnosing human and plant pathogens when combined with pre-amplification and lateral flow strips or fluorescence detection.

Conclusions:

  • Cas12c-DETECTOR represents a robust, sensitive, and precise nucleic acid detection platform.
  • The 5 acg-TG PAM recognition expands the utility of CRISPR-based detection systems.
  • Cas12c-DETECTOR shows significant potential for practical applications in diagnosing infectious diseases in humans and plants.