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Related Concept Videos

CRISPR/Cas9 Genome Editing01:28

CRISPR/Cas9 Genome Editing

73
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...
73
CRISPR01:59

CRISPR

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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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CRISPR and crRNAs02:53

CRISPR and crRNAs

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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.
The CRISPR-Cas system stores a copy of foreign DNA in the host genome and uses it to identify the foreign DNA upon reinfection. CRISPR-Cas has three different...
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Homologous Recombination02:31

Homologous Recombination

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The basic reaction of homologous recombination (HR) involves two chromatids that contain DNA sequences sharing a significant stretch of identity. One of these sequences uses a strand from another as a template to synthesize DNA in an enzyme-catalyzed reaction. The final product is a novel amalgamation of the two substrates. To ensure an accurate recombination of sequences, HR is restricted to the S and G2 phases of the cell cycle. At these stages, the DNA has been replicated already and the...
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Related Experiment Video

Updated: Jul 24, 2025

Enhanced Genome Editing with Cas9 Ribonucleoprotein in Diverse Cells and Organisms
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Enhanced Genome Editing with Cas9 Ribonucleoprotein in Diverse Cells and Organisms

Published on: May 25, 2018

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An engineered hypercompact CRISPR-Cas12f system with boosted gene-editing activity.

Tong Wu1,2, Chang Liu1, Siyuan Zou1,2

  • 1Department of Chemistry, The University of Chicago, Chicago, IL, USA.

Nature Chemical Biology
|July 3, 2023
PubMed
Summary
This summary is machine-generated.

Researchers engineered a smaller, more potent CRISPR-Cas enzyme, enAsCas12f, for precise gene editing. This enhanced system offers improved DNA cleavage activity and minimal off-target effects for genetic disorder treatments.

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Area of Science:

  • Molecular Biology
  • Biotechnology
  • Genetics

Background:

  • CRISPR-Cas systems are powerful tools for gene editing.
  • Compact CRISPR-Cas systems are desirable for therapeutic applications.
  • Existing systems often have limitations in editing activity and size.

Purpose of the Study:

  • To engineer a more potent and compact CRISPR-Cas enzyme.
  • To evaluate the gene-editing efficiency and specificity of the engineered system.
  • To elucidate the structural basis for the enzyme's activity.

Main Methods:

  • Protein engineering of AsCas12f to create enAsCas12f.
  • In vitro DNA cleavage assays.
  • Gene editing experiments in human cells.
  • Cryo-electron microscopy (cryo-EM) structural analysis.
  • Single guide RNA (sgRNA) engineering.

Main Results:

  • Engineered enAsCas12f is up to 11.3-fold more potent than wild-type AsCas12f.
  • enAsCas12f functions broadly in human cells, achieving up to 69.8% insertions and deletions.
  • High on-target activity with minimal off-target editing was observed.
  • Cryo-EM structure revealed dimerization-mediated substrate recognition.
  • Engineered sgRNA-v2 is shorter with comparable activity.

Conclusions:

  • The engineered hypercompact AsCas12f system enables robust and faithful gene editing.
  • enAsCas12f offers a promising tool for genetic disorder therapies.
  • Structural insights guide further optimization of CRISPR-Cas systems.