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相关概念视频

CRISPR01:59

CRISPR

57.5K
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...
57.5K
CRISPR/Cas9 Genome Editing01:28

CRISPR/Cas9 Genome Editing

1.7K
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...
18.7K
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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相关实验视频

Updated: Jan 15, 2026

Selection-dependent and Independent Generation of CRISPR/Cas9-mediated Gene Knockouts in Mammalian Cells
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Selection-dependent and Independent Generation of CRISPR/Cas9-mediated Gene Knockouts in Mammalian Cells

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通过CRISPR-Cas9评估温度依赖的DNA裂变

Alexa L Knight1, Jinping Luo2, George P Lisi1,3

  • 1Department of Molecular Biology, Cell Biology & Biochemistry, Brown University, Providence, RI, USA.

Bio-protocol
|October 13, 2025
PubMed
概括

这项研究提出了一种用于测量CRISPR-Cas9DNA裂变活性在不同温度的新协议. 这种方法有助于理解和设计Cas9酶用于各种应用,包括治疗用途.

关键词:
克里斯普尔-Cas9是什么意思在DNA裂变的过程中,细胞内核酶 (endonuclease) 是一种细胞内核酶.酵素酶是一种酶.热爱的人是热爱的人.

更多相关视频

Substrate Generation for Endonucleases of CRISPR/Cas Systems
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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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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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相关实验视频

Last Updated: Jan 15, 2026

Selection-dependent and Independent Generation of CRISPR/Cas9-mediated Gene Knockouts in Mammalian Cells
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Selection-dependent and Independent Generation of CRISPR/Cas9-mediated Gene Knockouts in Mammalian Cells

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Substrate Generation for Endonucleases of CRISPR/Cas Systems
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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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科学领域:

  • 生物化学 生物化学
  • 分子生物学分子生物学
  • 生物物理学的生物物理.

背景情况:

  • CRISPR-Cas9是一种具有治疗潜力的强大基因编辑工具,依赖于由RNA和PAM识别引导的DNA裂变.
  • Cas9激活涉及一个全性机制,影响其裂变活性和特异性.
  • 生物物理研究的目的是通过理解异质效应来设计增强的Cas效应器.

研究的目的:

  • 开发和验证一种协议,用于量化CRISPR-Cas9系统在不同温度范围内进行DNA裂变.
  • 为了能够评估中性和热性Cas9 ортолог的温度依赖功能.
  • 提供适用于各种Cas-RNP系统和基因组目标的多功能测试.

主要方法:

  • 调整已建立的体外DNA裂变试验,以纳入温度依赖的测量.
  • 利用来自热友生物 (例如,Geobacillus stearothermophilus) 和中友生物 (例如,SpCas9) 的Cas9.
  • 在不同生理温度调节下对DNA裂变的定性和定量评估.

主要成果:

  • 该协议成功量化了中性和热性Cas9系统的DNA裂变活性.
  • 它允许对功能温度范围进行评估,这对于极端友好的Cas系统至关重要.
  • 该测试可适应不同的基因组位置和原空间邻基因 (PAM) 要求.

结论:

  • 这个经过验证的协议提供了一种可靠的方法来评估CRISPR-Cas9.9的温度依赖DNA裂变.
  • 它通过描述它们的热活性配置文件,促进了增强的Cas效应器的工程设计.
  • 该试验的适应性使其成为各种CRISPR-Cas9研究和开发应用的宝贵工具.