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Related Concept Videos

Conservative Site-specific Recombination and Phase Variation02:53

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Because the DNA segments are cut and reorganized in a direction-specific manner, site-specific recombination has emerged as an efficient genetic engineering technique. Flippase and Cyclization recombinases or Flp and Cre, respectively, are two members of the tyrosine recombinase family derived from bacteriophages, that are used to mediate site-specific DNA insertions, deletions, and targeted expression of proteins in mammalian cell lines.
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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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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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Updated: Jun 5, 2025

CRISPR/Cas9-mediated Targeted Integration In Vivo Using a Homology-mediated End Joining-based Strategy
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CRISPR/Cas-Based Gene Editing Tools for Large DNA Fragment Integration.

Shuhan Yang1, Guang Hu2,3, Jianming Wang3

  • 1Institute of Nano Biomedicine and Engineering, Department of Instrument Science and Engineering, School of Electronic Information and Electrical Engineering, Shanghai Jiao Tong University, Shanghai 200240, China.

ACS Synthetic Biology
|December 16, 2024
PubMed
Summary

Efficiently inserting large DNA fragments into genomes is key for gene therapy. New CRISPR-based methods show promise for precise, large-scale gene editing, overcoming current limitations.

Keywords:
CRISPR/Casgenome editinghomology-directed repairlarge fragment knock-inrecombinase.transposase

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Area of Science:

  • Genomics
  • Molecular Biology
  • Biotechnology

Background:

  • Gene editing technologies enable precise genomic modification for understanding genetics and treating disorders.
  • Inserting large DNA fragments is crucial but faces challenges like inefficiency and off-target effects.

Purpose of the Study:

  • To review recent advancements (past 5 years) in integrating large DNA fragments into mammalian genomes.
  • To focus on CRISPR-related technologies for large fragment gene editing.
  • To explore potential applications in genetic engineering, gene therapy, and synthetic biology.

Main Methods:

  • Review of homology-directed repair (HDR) strategies.
  • Discussion of emerging CRISPR-transposase and CRISPR-recombinase techniques.
  • Analysis of methods for inserting kilobase-sized DNA fragments.

Main Results:

  • CRISPR-related technologies offer improved efficiency and precision for large DNA fragment integration.
  • Homology-directed repair, CRISPR-transposase, and CRISPR-recombinase strategies show significant potential.
  • These methods aim to overcome limitations of existing gene editing tools.

Conclusions:

  • Advancements in large fragment gene editing, particularly CRISPR-based methods, hold revolutionary potential for gene therapy.
  • Further development is needed to address challenges and facilitate broader application.
  • Improved tools will advance genetic engineering, gene therapy for complex diseases, and synthetic biology.