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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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CRISPR01:59

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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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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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A Rapid and Facile Pipeline for Generating Genomic Point Mutants in C. elegans Using CRISPR/Cas9 Ribonucleoproteins
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Efficient and Heritable Targeted Mutagenesis in Mosses Using the CRISPR/Cas9 System.

Toshihisa Nomura1, Tetsuya Sakurai2,3, Yuriko Osakabe2,4

  • 1RIKEN Center for Sustainable Resource Science, 1-7-22 Suehiro, Tsurumi, Yokohama, 230-0045 Japan toshihisa.nomura@riken.jp sakaki@riken.jp.

Plant & Cell Physiology
|December 18, 2016
PubMed
Summary

Researchers successfully applied the CRISPR/Cas9 gene editing system to mosses, enabling targeted mutations for studying plant evolution and bryology. This method efficiently modifies genes in moss protoplasts, accelerating research in non-model organisms.

Keywords:
BryophyteCRISPR/Cas9 systemGenome editingPhyscomitrella patensRNA-guided nucleasesScopelophila cataractae

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Area of Science:

  • Plant Biology
  • Genetics
  • Molecular Biology

Background:

  • The clustered regularly interspaced short palindromic repeat (CRISPR)/CRISPR-associated nuclease 9 (Cas9) system has revolutionized genome editing across various organisms.
  • Its application in non-model plant species, particularly bryophytes, remains less explored.
  • Mosses like Scopelophila cataractae (heavy metal tolerant) and Physcomitrella patens offer unique opportunities for evolutionary and ecological studies.

Purpose of the Study:

  • To establish and validate the CRISPR/Cas9 system for targeted genome modification in the non-model moss Scopelophila cataractae and the model moss Physcomitrella patens.
  • To assess the efficiency of CRISPR/Cas9 in generating phenotypic changes and various mutation types (insertions, deletions) in mosses.
  • To demonstrate the capability of multiplex gene editing and large deletion introduction using this system in mosses.

Main Methods:

  • Regeneration of moss plants from protoplasts.
  • Expression of single-chain guide RNA (sgRNA) and Cas9 nuclease within protoplasts.
  • Targeted mutagenesis via CRISPR/Cas9 system delivery into protoplasts.
  • Analysis of phenotypic changes and DNA sequencing to identify mutations.

Main Results:

  • Successful introduction of the CRISPR/Cas9 system into Scopelophila cataractae and Physcomitrella patens.
  • Achieved an acquisition rate of 45-69% for strains with phenotypic changes linked to target genes.
  • Observed various insertion and deletion mutations, including large deletions (approximately 3 kbp), and demonstrated multiplex gene editing capability.

Conclusions:

  • The CRISPR/Cas9 system is an effective tool for targeted genome modification in mosses.
  • This method accelerates research in bryology and the study of land plant evolution by enabling efficient genetic manipulation in non-model organisms.
  • The demonstrated efficiency and versatility of CRISPR/Cas9 in mosses open new avenues for functional genomics in bryophytes.