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相关概念视频

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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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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RNA Interference01:23

RNA Interference

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RNA interference (RNAi) is a process in which a small non-coding RNA molecule blocks the post-transcriptional expression of a gene by binding to its messenger RNA (mRNA) and preventing the protein from being translated.
This process occurs naturally in cells, often through the activity of genomically-encoded microRNAs. Researchers can take advantage of this mechanism by introducing synthetic RNAs to deactivate specific genes for research or therapeutic purposes. For example, RNAi could be used...
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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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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.
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Experimental RNAi02:15

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RNA interference (RNAi) is a cellular mechanism that inhibits gene expression by suppressing its transcription or activating the RNA degradation process. The mechanism was discovered by Andrew Fire and Craig Mello in 1998 in plants. Today, it is observed in almost all eukaryotes, including protozoa, flies, nematodes, insects, parasites, and mammals. This precise cellular mechanism of gene silencing has been developed into a technique that provides an efficient way to identify and determine the...
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相关实验视频

Updated: Jun 8, 2025

CIRCLE-Seq for Interrogation of Off-Target Gene Editing
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CIRCLE-Seq for Interrogation of Off-Target Gene Editing

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种子序列在CRISPR干扰系统中调解了目标外活动.

Neha Rohatgi1, Jean-Philippe Fortin2, Ted Lau3

  • 1Genentech Computational Sciences, Genentech, 1 DNA Way, South San Francisco, CA 94080, USA; Roche Informatics, Hoffman-La Roche Canada, 7070 Mississauga Road, Mississauga, ON, Canada.

Cell genomics
|November 7, 2024
PubMed
概括

克里斯普尔干扰 (CRISPRi) 系统可以导致广泛的基因沉默非目标效应. 这些意外影响主要是由于sgRNA和基因组DNA之间的种子序列互补性.

关键词:
在CRISPR中激活CRISPR.这就是CRISPR干扰的原因.PAM-近端种子序列的种子序列目标之外的活动活动.

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科学领域:

  • 分子生物学分子生物学
  • 遗传学 是一个遗传学.
  • 功能性基因组学 功能性基因组学

背景情况:

  • 克里斯普尔干扰 (CRISPRi) 是研究中基因沉默的一个关键技术.
  • 缺乏对CRISPRi目标外活动的系统调查.

研究的目的:

  • 为了研究CRISPRi系统的全基因组非目标活性.
  • 了解非目标效应对基因表达的影响.

主要方法:

  • 利用了一个全基因组的CRISPRi-Cas9单导向RNA (sgRNA) 库.
  • 分析了目标外活动及其对基因表达的影响.

主要成果:

  • 在CRISPRi中,非目标效应是普遍存在的,直接和间接地影响基因表达.
  • 大多数非目标与种子序列 (sgRNA间隔器的3'半) 与PAM近位基因组区域的互补性有关.
  • 目标外结合稳定性受到种子序列长度和不匹配容忍度的影响.

结论:

  • 在功能基因组学中,CRISPRi的目标外活动是一个重要的考虑因素.
  • 了解种子序列相互作用对于预测和减轻CRISPRi非目标效应至关重要.