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Related Concept Videos

CRISPR/Cas9 Genome Editing01:28

CRISPR/Cas9 Genome Editing

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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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CRISPR and crRNAs02:53

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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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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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Conservative Site-specific Recombination and Phase Variation02:53

Conservative Site-specific Recombination and Phase Variation

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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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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 22, 2025

A Rapid and Facile Pipeline for Generating Genomic Point Mutants in C. elegans Using CRISPR/Cas9 Ribonucleoproteins
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Highly efficient CRISPR-mediated gene editing in a rotifer.

Haiyang Feng1, Gemma Bavister1, Kristin E Gribble1

  • 1Josephine Bay Paul Center for Comparative Molecular Biology and Evolution, Marine Biological Laboratory, Woods Hole, Massachusetts, United States of America.

Plos Biology
|July 21, 2023
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Summary

Researchers developed a new CRISPR gene editing protocol for rotifers, enabling efficient knockout and knock-in mutations. This advancement significantly enhances rotifers as a model organism for studying gene function and biological discovery.

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CRISPR/Cas9 Editing of the C. elegans rbm-3.2 Gene using the dpy-10 Co-CRISPR Screening Marker and Assembled Ribonucleoprotein Complexes.
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Efficient Production and Identification of CRISPR/Cas9-generated Gene Knockouts in the Model System Danio rerio
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CRISPR/Cas9 Editing of the C. elegans rbm-3.2 Gene using the dpy-10 Co-CRISPR Screening Marker and Assembled Ribonucleoprotein Complexes.
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Efficient Production and Identification of CRISPR/Cas9-generated Gene Knockouts in the Model System Danio rerio
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Area of Science:

  • * Molecular Biology
  • * Developmental Biology
  • * Genetics

Background:

  • * Rotifers are valuable model organisms for studying microevolution, ecology, and toxicology.
  • * Previous limitations in gene editing tools hindered genotype-phenotype linkage in rotifers.
  • * Advancing rotifer genetics is crucial for dissecting molecular mechanisms of various biological processes.

Purpose of the Study:

  • * To develop a highly efficient, transgenerational CRISPR-mediated gene editing protocol for the rotifer Brachionus manjavacas.
  • * To establish a method for creating knockout and knock-in mutants in rotifers.
  • * To facilitate the creation of reporter lines and advance rotifers as a model system.

Main Methods:

  • * Microinjection of Cas9 protein and synthetic single-guide RNA into the vitellaria of young amictic female rotifers.
  • * CRISPR-mediated knockout of the developmental gene vasa and the DNA mismatch repair gene mlh3.
  • * CRISPR-mediated knock-in of a stop codon cassette into the mlh3 locus.

Main Results:

  • * Over 50% of injected mothers survived and produced offspring with CRISPR-induced mutations.
  • * Most offspring and subsequent generations carried at least one mutation, with many showing biallelic mutations.
  • * Successful knock-in of a stop codon cassette in the mlh3 locus was achieved in F2 offspring.

Conclusions:

  • * The developed protocol enables efficient, transgenerational CRISPR/Cas9 knockout and knock-in editing in rotifers.
  • * This method significantly advances the utility of rotifers as a model system for biological research.
  • * The protocol facilitates genotype-phenotype linkage and the dissection of molecular mechanisms in rotifers.