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CRISPR/Cas9 Genome Editing01:28

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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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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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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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Biosynthesis in bacteria is a fundamental anabolic process that generates essential macromolecules, including proteins, nucleic acids, lipids, and polysaccharides. These macromolecules are critical for cellular growth, replication, and function. The process is tightly regulated and energetically linked to catabolic pathways to ensure optimal resource utilization.Biosynthetic pathways begin with precursor metabolites such as pyruvate, acetyl-CoA, and glucose-6-phosphate derived from glycolysis,...
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Structure of PeptidoglycanPeptidoglycan is a vital structural component of the bacterial cell wall, providing mechanical strength and shape to the cell. It consists of repeating units of two sugars—N-acetylglucosamine (NAG) and N-acetylmuramic acid (NAM)—linked by β-1,4 glycosidic bonds. These sugar chains are cross-linked by short peptide chains, forming a mesh-like polymer that surrounds the bacterial plasma membrane.Cytoplasmic Phase – Precursor SynthesisPeptidoglycan...
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Engineering Modular Polyketide Biosynthesis in Streptomyces Using CRISPR/Cas: A Practical Guide.

Jean-Malo Massicard1, Li Su1, Christophe Jacob2

  • 1Molecular and Structural Enzymology Group, UMR 7365 CNRS-UL IMoPA, Lorraine University, Faculté de médecine, Batiment Biopôle, Vandœuvre-lès-Nancy Cedex, France.

Methods in Molecular Biology (Clifton, N.J.)
|May 6, 2022
PubMed
Summary
This summary is machine-generated.

CRISPR/Cas gene editing is now adaptable for Streptomyces bacteria to precisely alter polyketide natural product biosynthesis. This guide aids researchers in applying this powerful tool for medical and agricultural applications.

Keywords:
CRISPR/CasGenome editingPolyketide biosynthesisStreptomyces

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Area of Science:

  • Microbiology
  • Molecular Biology
  • Biochemistry

Background:

  • The CRISPR/Cas system is a versatile gene-editing tool successfully applied across various organisms.
  • Streptomyces bacteria are crucial producers of polyketide natural products with significant medical and agricultural value.
  • Rational modification of polyketide biosynthesis pathways is essential for optimizing natural product yields and properties.

Purpose of the Study:

  • To provide a practical guide for utilizing the CRISPR/Cas system in Streptomyces.
  • To facilitate the manipulation of polyketide biosynthesis pathways in these bacteria.
  • To assist researchers in adapting CRISPR/Cas technology for Streptomyces polyketide production.

Main Methods:

  • Adaptation of the CRISPR/Cas system for use in Streptomyces species.
  • Strategic design of genetic engineering components for pathway manipulation.
  • Detailed troubleshooting for common experimental challenges.

Main Results:

  • Demonstration of CRISPR/Cas system applicability in Streptomyces for genetic engineering.
  • Successful modification of polyketide biosynthetic pathways.
  • Establishment of a reliable protocol for CRISPR/Cas-mediated manipulation.

Conclusions:

  • The CRISPR/Cas system offers a powerful and adaptable approach for engineering Streptomyces.
  • This methodology enables precise manipulation of polyketide biosynthesis pathways.
  • The provided guide empowers researchers to leverage CRISPR/Cas for advancing natural product discovery and production.