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相关概念视频

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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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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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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.
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相关实验视频

Updated: Mar 23, 2026

Gene Knock-in by CRISPR/Cas9 and Cell Sorting in Macrophage and T Cell Lines
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使用CRISPR/Cas9对活细胞进行可编程RNA追踪

David A Nelles1, Mark Y Fang2, Mitchell R O'Connell3

  • 1Department of Cellular and Molecular Medicine and Institute for Genomic Medicine, University of California, San Diego, La Jolla, CA 92037, USA; Materials Science and Engineering Graduate Program, University of California, San Diego, La Jolla, CA 92093, USA.

Cell
|March 22, 2016
PubMed
概括

研究人员开发了一种新的CRISPR/Cas9工具 (RCas9),在不需要遗传标签的情况下追踪活细胞中的RNA. 这种针对RNA的Cas9系统可以实时对内源信使RNA (mRNA) 进行可编程可视化.

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科学领域:

  • 分子生物学
  • 遗传学
  • 生物技术

背景情况:

  • 来自Streptococcus pyogenes的CRISPR/Cas9系统在基因组编辑方面非常强大.
  • 目前的RNA向方法通常需要外源标签,限制了灵活性.
  • 需要一个可编程的RNA向工具来进行内源RNA分析和成像.

研究的目的:

  • 展示可编程RNA向的核酶非活性CRISPR/Cas9系统.
  • 在没有遗传标签的活细胞中实现内源性RNA跟踪和成像.
  • 调查mRNA局部化和贩运动态.

主要方法:

  • 使用用于RNA结合的非核酶活性的Streptococcus pyogenes CRISPR/Cas9.
  • 使用指导RNA (sgRNA) 来编程RCas9针对特定的信使RNA (mRNA).
  • 使用显微镜和现场光杂交观察RCas9局部化和mRNA积累.

主要成果:

  • 核局部化RCas9在结合 sgRNA 向 mRNA 时被输出到细胞质.
  • 在RNA颗粒中观察到ACTB,CCNA2和TFRCmRNA的积累.
  • 证明了ACTBmRNA的时间解决跟踪到压力颗粒,揭示了贩运动态.

结论:

  • RCas9提供了一个可编程的,无标签的方法来追踪活细胞中的内源RNA.
  • 这种向RNA的Cas9系统有助于研究mRNA的局部化和动态.
  • 这些发现为RNA生物学研究建立了一个类似于基因组CRISPR工具的新工具.