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Related Concept Videos

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Genome editing technologies allow scientists to modify an organism’s DNA via the addition, removal, or rearrangement of genetic material at specific genomic locations. These types of techniques could potentially be used to cure genetic disorders such as hemophilia and sickle cell anemia. One popular and widely used DNA-editing research tool that could lead to safe and effective cures for genetic disorders is the CRISPR-Cas9 system. CRISPR-Cas9 stands for Clustered Regularly Interspaced...
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Bacteria and archaea are susceptible to viral infections just like eukaryotes; therefore, they have developed a unique adaptive immune system to protect themselves. Clustered regularly interspaced short palindromic repeats and CRISPR-associated proteins (CRISPR-Cas) are present in more than 45% of known bacteria and 90% of known archaea.
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Updated: Jun 6, 2025

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DNA-guided CRISPR/Cas12 for RNA targeting.

Carlos Orosco1, Santosh R Rananaware1, Boyu Huang1

  • 1Department of Chemical Engineering, University of Florida, Gainesville, FL, USA.

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|November 28, 2024
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Summary

Researchers developed ΨDNA, a DNA-based guide for Cas12 enzymes, enabling precise RNA targeting for diagnostics and cellular RNA modulation. This innovation bypasses traditional RNA guides for enhanced CRISPR applications.

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Area of Science:

  • Molecular Biology
  • Biotechnology
  • Genetics

Background:

  • CRISPR-Cas nucleases are powerful tools for genome editing, RNA editing, and diagnostics.
  • Current CRISPR systems primarily rely on RNA-guided mechanisms, limiting their versatility.

Purpose of the Study:

  • To introduce ΨDNA, a novel DNA-based guide for Cas12 enzymes, for efficient and specific RNA targeting.
  • To explore the potential of ΨDNA in diagnostic applications and cellular RNA modulation.

Main Methods:

  • Engineered ΨDNA molecules that mimic crRNA in a reverse orientation for Cas12-RNA assembly.
  • Tested ΨDNA for RNA sensing capabilities, including short and long RNA detection.
  • Evaluated ΨDNA's performance in detecting Hepatitis C Virus (HCV) RNA in clinical samples.
  • Investigated ΨDNA's ability to guide Cas12 enzymes for RNA targeting within cells, including mRNA degradation and multiplex knockdown.

Main Results:

  • ΨDNA demonstrated stable Cas12-RNA assembly and activated trans-cleavage activity without RNA components.
  • ΨDNAs showed high efficacy in sensing both short and long RNA targets.
  • Achieved 100% accuracy in detecting HCV RNA from clinical samples.
  • Successfully guided Cas12 enzymes for RNA targeting in cellular environments, leading to enhanced mRNA degradation and multiplex RNA transcript knockdown.

Conclusions:

  • ΨDNA represents a robust DNA-based alternative to traditional RNA guides for CRISPR-Cas12 systems.
  • This technology significantly expands the potential of CRISPR-Cas12 for advanced diagnostic tools.
  • ΨDNA enables targeted RNA modulation within cellular contexts, offering new therapeutic and research avenues.