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Related Concept Videos

CRISPR01:59

CRISPR

51.0K
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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Reporter Genes02:11

Reporter Genes

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Reporter genes are a type of protein-coding gene that are often tagged to a gene of interest. Once inside a target cell, reporter genes usually produce visually identifiable characteristics like fluorescence and luminescence when expressed along with the gene of interest. Thus, reporter genes “report” the presence or absence of genes of interest in an organism, determine the gene expression pattern, or track the physical location of a DNA segment or protein in the cell.
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Related Experiment Video

Updated: Jul 4, 2025

Using a Fluorescent PCR-capillary Gel Electrophoresis Technique to Genotype CRISPR/Cas9-mediated Knockout Mutants in a High-throughput Format
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Using a Fluorescent PCR-capillary Gel Electrophoresis Technique to Genotype CRISPR/Cas9-mediated Knockout Mutants in a High-throughput Format

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Fluorogenic CRISPR for genomic DNA imaging.

Zhongxuan Zhang1,2,3,4, Xiaoxiao Rong1,5, Tianjin Xie1,6

  • 1Beijing Institute of Life Sciences, Chinese Academy of Science, 100101, Beijing, China.

Nature Communications
|January 31, 2024
PubMed
Summary
This summary is machine-generated.

Fluorogenic CRISPR (fCRISPR) enables high-contrast imaging of genomic DNA in living cells. This novel CRISPR tool overcomes background noise, allowing sensitive tracking of chromosome dynamics and DNA repair in real time.

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A Rapid and Facile Pipeline for Generating Genomic Point Mutants in C. elegans Using CRISPR/Cas9 Ribonucleoproteins
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Area of Science:

  • Molecular Biology
  • Genetics
  • Cell Biology

Background:

  • Genomic DNA displays significant heterogeneity in nuclear dynamics, structure, and function.
  • CRISPR-based imaging allows visualization of genomic loci in living cells.
  • Conventional CRISPR tools suffer from high background and nonspecific signals due to constitutive fluorescent protein expression.

Purpose of the Study:

  • To develop a novel CRISPR-based imaging tool with improved signal-to-noise ratio and sensitivity.
  • To overcome limitations of existing CRISPR imaging techniques, such as high background and nonspecific signals.
  • To enable high-contrast, real-time imaging of genomic DNA dynamics and repair processes.

Main Methods:

  • Construction of fluorogenic CRISPR (fCRISPR) utilizing dCas9, an engineered sgRNA, and a fluorogenic protein.
  • Incorporation of RNA hairpins into sgRNA to trigger fluorogenic protein activation upon binding.
  • Formation of dCas9: sgRNA: fluorogenic protein ternary complexes for targeted DNA imaging.

Main Results:

  • fCRISPR achieved high signal-to-noise ratio and sensitivity for imaging diverse genomic DNA in human cells.
  • The system successfully tracked chromosome dynamics and length.
  • Real-time monitoring of DNA double-strand breaks (DSBs) and their repair was enabled by fCRISPR.

Conclusions:

  • fCRISPR provides a high-contrast and sensitive platform for imaging genomic loci in living cells.
  • This technology overcomes the limitations of conventional CRISPR imaging, offering improved specificity and reduced background.
  • fCRISPR facilitates advanced studies on chromosome dynamics and DNA repair mechanisms.