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相关概念视频

CRISPR01:59

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 Short...
Homologous Recombination02:31

Homologous Recombination

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...
Conservative Site-specific Recombination and Phase Variation02:53

Conservative Site-specific Recombination and Phase Variation

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...
CRISPR/Cas9 Genome Editing01:28

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...

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相关实验视频

Updated: Jun 23, 2026

Substrate Generation for Endonucleases of CRISPR/Cas Systems
11:53

Substrate Generation for Endonucleases of CRISPR/Cas Systems

Published on: September 8, 2012

27.4K

通过多重CRISPR查来定义E3酶-基质关系.

Richard T Timms1,2, Elijah L Mena1, Yumei Leng1

  • 1Department of Genetics, Harvard Medical School, Division of Genetics, Brigham asnd Women's Hospital, Howard Hughes Medical Institute, Boston, MA, USA.

Nature cell biology
|September 22, 2023
PubMed
概括
此摘要是机器生成的。

研究人员开发了一种多重CRISPR选平台,以识别E3无素合酶的基质. 这种高通量方法加快了对ubiquitin-proteasome系统的理解.

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Investigation of Genetic Dependencies Using CRISPR-Cas9-based Competition Assays
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Investigation of Genetic Dependencies Using CRISPR-Cas9-based Competition Assays

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Pooled CRISPR-Based Genetic Screens in Mammalian Cells
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Pooled CRISPR-Based Genetic Screens in Mammalian Cells

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相关实验视频

Last Updated: Jun 23, 2026

Substrate Generation for Endonucleases of CRISPR/Cas Systems
11:53

Substrate Generation for Endonucleases of CRISPR/Cas Systems

Published on: September 8, 2012

27.4K
Investigation of Genetic Dependencies Using CRISPR-Cas9-based Competition Assays
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Investigation of Genetic Dependencies Using CRISPR-Cas9-based Competition Assays

Published on: January 7, 2019

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Pooled CRISPR-Based Genetic Screens in Mammalian Cells
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科学领域:

  • 分子生物学分子生物学
  • 遗传学 遗传学 是一个
  • 生物化学 生物化学

背景情况:

  • 无素-蛋白酶体系统的特异性依赖于E3无素连接酶,但它们的基质和识别动机 (退基) 往往是未知的.
  • 现有的E3酶基质分配方法耗时且产量低.

研究的目的:

  • 开发一个可扩展和高吞吐量平台,用于将E3无素连接酶分配给它们的相关基质.
  • 确定各种E3结合酶的新型基质识别途径和降解动机.

主要方法:

  • 开发一个多重CRISPR选平台,使多个选器能够同时执行.
  • 在单个实验中,涉及~100个CRISPR屏幕的原理验证屏幕.
  • 与位点和突变发生的整合,以确定特定的降解动机.

主要成果:

  • 成功完善已知的C-降解途径,并确定了一条新的途径,涉及Cul2FEM1B针对C-终端.
  • 已确定多个E3酶的基质,包括Cul1FBXO38,Cul2APPBP2,Cul3GAN,Cul3KLHL8,Cul3KLHL9/13,以及Cul3KLHL15.
  • 证明了平台与各种蛋白质基质稳定性的兼容性,以及其赋予降解动机的能力.

结论:

  • 多重CRISPR选是一种强大的工具,用于大规模的E3酶基质分配.
  • 这种方法显著加快了E3酶特异性和降级识别的表征.
  • 这些发现将有助于更好地理解无素-蛋白酶体系统调节的理解.