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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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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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Recombinant DNA technology called transgenesis is often used to add a foreign gene or remove a detrimental gene from an organism. Such genetically modified organisms are called transgenic organisms.
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Efficient plant genome engineering using a probiotic sourced CRISPR-Cas9 system.

Zhaohui Zhong1,2, Guanqing Liu3,4,5, Zhongjie Tang1

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Researchers developed a new CRISPR-Cas9 genome editing system from Lactobacillus rhamnosus (LrCas9) for plants. This efficient tool surpasses existing systems in editing various crops like rice and wheat.

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Area of Science:

  • Molecular Biology
  • Plant Science
  • Biotechnology

Background:

  • Streptococcus pyogenes Cas9 (SpCas9) is the most common CRISPR-Cas genome editing system.
  • SpCas9 is derived from a human pathogen, prompting the search for alternative systems.
  • There is a need for efficient and versatile genome engineering tools in plant biotechnology.

Purpose of the Study:

  • To establish an efficient plant genome engineering system using CRISPR-Cas9 from Lactobacillus rhamnosus (LrCas9).
  • To validate the 5'-NGAAA-3' Protospacer Adjacent Motif (PAM) of LrCas9.
  • To demonstrate the editing efficiency and specificity of LrCas9 in various plant species and applications.

Main Methods:

  • In silico data mining to identify potential CRISPR-Cas9 systems.
  • Bacterial PAM depletion assay to confirm the LrCas9 PAM sequence.
  • Genome editing experiments in rice, wheat, tomato, and Larix cells.
  • Development of LrCas9-derived base editors and CRISPR interference/activation systems.

Main Results:

  • LrCas9 exhibits exceptional editing efficiency in rice, wheat, tomato, and Larix cells, outperforming LbCas12a, SpCas9-NG, and SpRY.
  • Multiplexed gene knockout and gene knockdown were achieved in stable rice lines using LrCas9.
  • High specificity of LrCas9 was demonstrated through targeted promoter deletion and coding sequence editing.
  • LrCas9-derived cytosine and adenine base editors were successfully developed.
  • Efficient CRISPR interference and activation systems were created by leveraging LrCas9's A/T-rich PAM targeting.

Conclusions:

  • CRISPR-LrCas9 is an efficient and user-friendly genome engineering tool for plants.
  • LrCas9 offers advantages over existing systems, including broader PAM targeting and higher efficiency.
  • This system expands genome editing capabilities in crops and has potential for diverse applications.