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相关概念视频

CRISPR01:59

CRISPR

50.8K
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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Homologous Recombination02:31

Homologous Recombination

50.5K
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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CRISPR and crRNAs02:53

CRISPR and crRNAs

17.0K
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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Cas9 RNP 物理化学分析用于增强CRISPR-AuNP组件和功能.

Daniel D Lane1, Karthikeya S V Gottimukkala1,2, Rachel A Cunningham1,2

  • 1Translational Science and Therapeutics Division, Fred Hutchinson Cancer Center, Seattle, WA, USA.

bioRxiv : the preprint server for biology
|April 15, 2024
PubMed
概括

研究人员开发了改进的CRISPR-gold纳米颗粒,用于在血液形成干细胞中的体内基因编辑. 这种方法增强了CRISPR-Cas9的传递和活性,有可能克服目前血液疾病的ex vivo疗法的局限性.

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科学领域:

  • 基因编辑和纳米医学用于血液学疾病.

背景情况:

  • 目前针对β血症和状细胞贫血的CRISPR疗法需要对造血干细胞和原生细胞 (HSPC) 进行ex vivo操纵,涉及有毒电穿孔和化疗调节.
  • 通过纳米载体进行CRISPR基因编辑工具的体内输送提供了一个有希望的替代方案,以减轻复杂性和毒性,但在克服HSPC限制因素方面面临挑战.
  • 以前的CRISPR-金纳米粒子 (CRISPR-AuNP) 系统显示出不一致的Cas9活性,这是由于加载过程中RNA的不稳定性.

研究的目的:

  • 开发一种携带CRISPR的黄金纳米粒子 (CRISPR-AuNP),以便在体内有效地将CRISPR-Cas核糖蛋白复合体 (RNP) 输送到HSPC中.
  • 为了克服最初的CRISPR-AuNP配方中观察到的Cas9活性和负载不稳定的局限性.
  • 为了提高纳米颗粒的稳定性和潜在的 in vivo 管理的特性.

主要方法:

  • 在加载到金纳米粒子 (AuNP) 之前,Cas9和Cas12a RNP复合物的预制.
  • 优化RNP加载化学和条件,以改善粒子结合.
  • 用PEGylation修改纳米粒子外层,以提高稳定性和表面性能.

主要成果:

  • 在不影响活动的情况下实现了39.6 ± 7.0 Cas9 RNP/AuNP的显著负载,并且Cas12a RNP/AuNP负载增加了10倍.
  • 开发了第二代CRISPR-AuNP,其稳定性得到了改善,并且具有适合体内使用的水友性中性表面.
  • 通过初级人类HSPCs证明了第二代CRISPR-AuNP的高体外吸收率 (72.5 ± 7.37%),尽管内体积累限制了基因编辑效率.

结论:

  • 预制RNP和优化加载化学对于通过黄金纳米粒子在体内有效传递CRISPR-Cas9和Cas12a基因编辑系统至关重要.
  • 开发的CRISPR-AuNP纳米配方显示了HSPC体内体内基因编辑的潜力,但需要进一步改进,以提高内体逃生和基因编辑效率.