関連する実験動画

Updated: Feb 14, 2026

Preparing and Injecting Embryos of Culex Mosquitoes to Generate Null Mutations using CRISPR/Cas9

Preparing and Injecting Embryos of Culex Mosquitoes to Generate Null Mutations using CRISPR/Cas9

Published on: September 10, 2020

CRISPR/Cas9の運動メカニズムによって明らかになった機能的洞察

Austin T Raper¹, Anthony A Stephenson¹, Zucai Suo¹

¹Department of Chemistry and Biochemistry, ‡Ohio State Biochemistry Program, and §The James Comprehensive Cancer Center, The Ohio State University , Columbus, Ohio 43210, United States.

Journal of the American Chemical Society

|February 15, 2018

まとめ

この研究は,Streptococcus pyogenes Cas9の運動メカニズムを定義し,ゆっくりとしたDNA産物放出が単一ターンオーバー酵素であることを明らかにしました. Cas9の運動を理解することは遺伝子編集技術の進歩に不可欠です

科学分野:

生物化学
分子生物学
遺伝学

背景:

プロカリオットの適応免疫は,遺伝子工学におけるCRISPR-Cas9の広範な使用につながった.
Cas9の運動メカニズムとDNA分裂の詳細は十分に理解されていません.

研究の目的:

ガイドRNA結合からDNA分裂と産物放出に至るStreptococcus pyogenes Cas9の運動メカニズムを確立する.
Cas9内のHNHとRuvC核酵素活性調節の運動的基礎を解明する.

主な方法:

ストレプトコッカス・ピオゲネスCas9の運動分析
DNA分裂へのHNHとRuvC核酵素の貢献を同時に測定する.

主要な成果:

DNA結合は最初のターンオーバーで速度を制限し,三元複合体からの分裂は迅速である.
非常に遅いDNA産物放出により,Cas9は単回転活性に制限されます.
HNHドメインはRuvC核酵素の活性を構成的に調節する.

結論:

Cas9の重要な動力学的および機能的な詳細を提供します.
遺伝子編集の実験や標的外DNAの分裂に関する研究を
バイオテクノロジーのツールとしてCas9の継続的な開発を支援する.

さらに関連する動画

Using CRISPR/Cas9 to Knock Out GM-CSF in CAR-T Cells

Using CRISPR/Cas9 to Knock Out GM-CSF in CAR-T Cells

Published on: July 22, 2019

Construction of Homozygous Mutants of Migratory Locust Using CRISPR/Cas9 Technology

Construction of Homozygous Mutants of Migratory Locust Using CRISPR/Cas9 Technology

Published on: March 16, 2022

関連する概念動画

CRISPR

CRISPR

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...

CRISPR/Cas9 Genome Editing

CRISPR/Cas9 Genome Editing

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...

CRISPR and crRNAs

CRISPR and crRNAs

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...

Mechanical Protein Functions

Mechanical Protein Functions

Proteins perform many mechanical functions in a cell. These proteins can be classified into two general categories- proteins that generate mechanical forces and proteins that are subjected to mechanical forces. Proteins providing mechanical support to the structure of the cell, such as keratin, are subjected to mechanical force, whereas proteins involved in cell movement and transport of molecules across cell membranes, such as an ion pump, are examples of generating mechanical force.

E2 Reaction: Kinetics and Mechanism

E2 Reaction: Kinetics and Mechanism

SN2 substitutions and E2 eliminations of alkyl halides proceed via a concerted pathway. While the nucleophile attacks the alpha carbon in SN2 reactions, it functions as a strong base and abstracts a beta hydrogen in the E2 mechanism. The rate-limiting transition state in E2 elimination reactions is characterized by partially broken carbon–hydrogen and carbon–halogen bonds and a partially formed pi bond between the alpha and beta carbons. The beta hydrogen and halide are eliminated...

E1 Reaction: Kinetics and Mechanism

E1 Reaction: Kinetics and Mechanism

Here, in contrast to the E2 reaction mechanism, we delve into the aspects of the E1 reaction mechanism, which has two steps: rate-limiting loss of the leaving group and abstraction of the beta hydrogen by a weak base. Typically, the experimental proof for the E1 mechanism is via kinetic studies or isotope studies. While the former demonstrates the first-order kinetics—the dependence of the reaction solely on substrate concentration—the latter proves the abstraction of hydrogen only...