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CRISPR/Cas9 Genome Editing01:28

CRISPR/Cas9 Genome Editing

1.5K
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

CRISPR and crRNAs

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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.
The recognition sites for Cre recombinase called LoxP...
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Updated: Dec 28, 2025

Genome Editing in Mammalian Cell Lines using CRISPR-Cas
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精密なCRISPR適応のためのプリスペーサーの選択的なロードと処理

Sungchul Kim1, Luuk Loeff2, Sabina Colombo2

  • 1Kavli Institute of Nanoscience, Department of Bionanoscience, Delft University of Technology, Delft, The Netherlands. sungchulkim.kr@gmail.com.

Nature
|February 21, 2020
PubMed
まとめ

CRISPR-CasシステムはCas1-Cas2複合体を用いて外来DNAの記憶を取得します この研究では,Cas1-Cas2がDNA断片を選択し,DnaQエクソヌクレアスを使ってそれらをカットし,プロカリオット免疫のための正しいスペーサー統合を確保する方法が明らかになりました.

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科学分野:

  • 分子生物学
  • 微生物学
  • 遺伝学

背景:

  • CRISPR-Cas免疫は,外来遺伝子要素に対するプロカリオットの防御を提供します.
  • Cas1-Cas2複合体は,スペーサー取得を通じて遺伝性免疫記憶の確立に不可欠である.
  • Cas1-Cas2によるプレスペーサー選択と統合のメカニズムは,まだ完全に理解されていません.

研究 の 目的:

  • Cas1-Cas2が外来DNAからプレスペーサー前駆体を選択するメカニズムを解明する.
  • 統合のためのプリスペーサーの処理に関与する酵素を特定する.
  • CRISPRの位置に統合する際に正しいスペーサーの方向性がどのように達成されるかを理解する.

主な方法:

  • 高解像度の単分子光顕微鏡
  • 様々な形態のCas1-Cas2によるDNA選択の分析 (ssDNA,部分複合体).
  • プレスペーサー成熟におけるDnaQエクソヌクレアスの識別

主要な成果:

  • Cas1-Cas2は,DNAの長さとプロトスペーサー隣接モチーフ (PAM) の存在に基づいてプレスペーサー前駆者を選択します.
  • DnaQエクソヌクレアゼは,選択された前駆体を成熟した,統合可能なプレスペーサーに処理する.
  • Cas1-Cas2はPAM配列を保護し,非対称なトリミングと正しいスペーサーの方向性を導きます.

結論:

  • この研究は,Cas1-Cas2によるプレスペーサー前駆体選択とPAMトリミングの調整メカニズムを示しています.
  • このプロセスは,適応免疫のためのCRISPRロシに機能的なスペーサーを正確に統合することを保証します.
  • この発見は,CRISPR-Casの適応免疫の分子メカニズムに関する重要な洞察を提供します.