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The CRISPR-Cas system serves as a bacterial defense mechanism against invading genetic elements such as viruses and plasmids, forming the foundation for its adaptation as a powerful genome-editing tool. Originally discovered in prokaryotes, this system has been repurposed to revolutionize genetic engineering across a wide range of organisms, including plants, animals, and humans. The core component, Cas9, is an endonuclease derived from Streptococcus pyogenes, capable of introducing...
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Plant-Based Biosensors for Detecting CRISPR-Mediated Genome Engineering.

Guoliang Yuan1,2, Md Mahmudul Hassan1,2,3, Tao Yao1,2

  • 1Biosciences Division, Oak Ridge National Laboratory, Oak Ridge, Tennessee 37831, United States.

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|December 8, 2021
PubMed
Summary

Researchers developed four real-time detection systems to identify active CRISPR/Cas gene editing tools, including CRISPR/Cas9 nuclease, base editing, prime editing, and CRISPRa, in plants. This innovation addresses safety concerns by enabling detection of unintended DNA changes.

Keywords:
CRISPRbiosensordetectiongenome editingtransient gene expression

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Area of Science:

  • Molecular Biology
  • Genetics
  • Biotechnology

Background:

  • CRISPR/Cas technology is a powerful tool for genome engineering.
  • Concerns exist regarding unintended DNA modifications from CRISPR gene editing.
  • A need for systems to detect active CRISPR/Cas tools in biological systems is crucial.

Purpose of the Study:

  • To develop and validate real-time detection systems for active CRISPR/Cas tools in plants.
  • To monitor genome editing and gene regulation activities.
  • To address safety concerns associated with CRISPR technologies.

Main Methods:

  • Development of four fluorescence-based molecular biosensors.
  • Detection of CRISPR/Cas9 nuclease, base editing, prime editing, and CRISPRa activities.
  • Application in transient expression (protoplast transformation, leaf infiltration) and stable transformation in plants (Arabidopsis, poplar, tobacco).

Main Results:

  • Successfully demonstrated real-time detection of active CRISPR/Cas tools.
  • Validated the system's efficacy across different CRISPR applications (nuclease, base editing, prime editing, CRISPRa).
  • Confirmed detection in various plant species and transformation methods.

Conclusions:

  • The developed biosensors provide an effective means to detect active CRISPR/Cas tools in plants.
  • This system aids in monitoring genome editing processes and ensuring safety.
  • Facilitates reliable application of CRISPR technologies in plant research and breeding.