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Related Concept Videos

CRISPR/Cas9 Genome Editing01:28

CRISPR/Cas9 Genome Editing

59
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...
59
CRISPR01:59

CRISPR

52.3K
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...
52.3K

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Related Experiment Video

Updated: Jul 19, 2025

Efficient Production and Identification of CRISPR/Cas9-generated Gene Knockouts in the Model System Danio rerio
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Optimized CRISPR/Cas9 system for gene knockout in chicken DF1 cells.

Kexin Zou1, Fang Wang1, Zechun Zhang1

  • 1College of Animal Science and Technology, Northwest A&F University, Yangling, Shaanxi 712100, China.

Poultry Science
|August 10, 2023
PubMed
Summary

CRISPR/Cas9 gene editing efficiency in chicken cells was improved using surrogate reporters. Dual sgRNAs increased fragment deletions, and amplicon sequencing proved more sensitive than T7E1 assays for evaluating editing outcomes.

Keywords:
CRISPRDF1 cellsT7E1amplicon sequencedual sgRNA

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Area of Science:

  • Molecular Biology
  • Genetics
  • Biotechnology

Background:

  • CRISPR technology's utility in poultry is limited by editing efficiency.
  • Understanding factors influencing CRISPR/Cas9 gene knockout efficiency in chicken cells is crucial.

Purpose of the Study:

  • To investigate factors affecting CRISPR/Cas9 gene knockout efficiency in chicken DF1 cells.
  • To compare genome editing evaluation methods and analyze indel profiles.

Main Methods:

  • Designed single or dual sgRNAs targeting specific gene exons (AMH, TGFBR2, PPARG).
  • Utilized sgRNA-CRISPR/Cas9 and reporter vectors in chicken DF1 cells.
  • Evaluated editing efficiency using T7E1 and amplicon sequencing assays.

Main Results:

  • Surrogate reporter systems enhanced editing efficiency, with increased indel percentages post-selection.
  • Amplicon sequencing showed higher sensitivity than T7E1 assays for detecting cleavage.
  • Dual sgRNAs increased fragment deletion ratios compared to single sgRNAs.

Conclusions:

  • Surrogate reporters effectively promote CRISPR/Cas9 editing efficiency in chicken cells.
  • Amplicon sequencing is a more sensitive method for evaluating CRISPR/Cas9 targeting efficiency than T7E1 assays.
  • Dual sgRNAs can enhance fragment deletions, offering a strategy for specific genomic modifications in poultry.