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相关概念视频

Single-Strand DNA Binding Proteins01:03

Single-Strand DNA Binding Proteins

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For successful DNA replication, the unwinding of double-stranded DNA must be accompanied by stabilization and protection of the separated single strands of the DNA. This crucial task is performed by single-strand DNA-binding (SSB) proteins. They bind to the DNA in a sequence-independent manner, which means that the nitrogenous bases of the DNA need not be present in a specific order for binding of SSB proteins to it. The binding of SSB proteins straightens single-stranded DNA (ssDNA) and makes...
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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...
16.7K
DNA Helicases00:55

DNA Helicases

21.1K
DNA unwinding helicase enzymes are a type of motor protein. Motor proteins can translocate along filaments or polymers using energy generated from ATP hydrolysis. Helicases are involved in all the important cellular processes where DNA unwinding is required, such as DNA replication, repair, recombination, and transcription. They are present in all living organisms, but vary in their structure, function, and mechanism of action. For example, in prokaryotes, DnaB helicase binds and translocates...
21.1K
Fixing Double-strand Breaks02:04

Fixing Double-strand Breaks

12.2K
The double-stranded structure of DNA has two major advantages. First, it serves as a safe repository of genetic information where one strand serves as the back-up in case the other strand is damaged. Second, the double-helical structure can be wrapped around proteins called histones to form nucleosomes, which can then be tightly wound to form chromosomes. This way, DNA chains up to 2 inches long can be contained within microscopic structures in a cell. A double-stranded break not only damages...
12.2K
Caspases01:24

Caspases

12.3K
Caspase, a family of cysteine proteases, serve as effectors in apoptosis. The ced3 gene in C.elegans was first identified to be involved in apoptosis. This gene encodes the ced-3 caspase that is similar to the interleukin-1-beta converting enzyme or ICE in mammals. In addition to apoptosis, caspases also function in the inflammatory response. Inflammatory caspases are essential in activating pro-inflammatory cytokines that recruit immune cells and block the replication of pathogens inside...
12.3K
Translesion DNA Polymerases02:10

Translesion DNA Polymerases

9.8K
Translesion (TLS) polymerases rescue stalled DNA polymerases at sites of damaged bases by replacing the replicative polymerase and installing a nucleotide across the damaged site. Doing so, TLS allows additional time for the cell to repair the damage before resuming regular DNA replication.
TLS polymerases are found in all three domains of life - archaea, bacteria, and eukaryotes. Of the different classes of TLS polymerases, members of the Y family are fitted with specialized structures that...
9.8K

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相关实验视频

Updated: Jun 4, 2025

Substrate Generation for Endonucleases of CRISPR/Cas Systems
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Substrate Generation for Endonucleases of CRISPR/Cas Systems

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在Cas12e中,ortologs开发了可变的结构元素,以促进dSDNA裂变.

Danyuan Li1,2, Shouyue Zhang3,4, Shuo Lin1,2

  • 1Beijing Frontier Research Center for Biological Structure, State Key Laboratory of Membrane Biology, School of Life Sciences, Tsinghua University, Beijing, 100084, China.

Nature communications
|December 31, 2024
PubMed
概括

研究人员探索了各种CRISPR-Cas系统,特别是Cas12e核酶,用于DNA操纵. 他们发现了新的Cas12e变体具有独特的特性,增强了基因组编辑工具.

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In Vitro Directed Evolution of a Restriction Endonuclease with More Stringent Specificity
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In Vitro Directed Evolution of a Restriction Endonuclease with More Stringent Specificity

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Parallel High Throughput Single Molecule Kinetic Assay for Site-Specific DNA Cleavage
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Parallel High Throughput Single Molecule Kinetic Assay for Site-Specific DNA Cleavage

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相关实验视频

Last Updated: Jun 4, 2025

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11:53

Substrate Generation for Endonucleases of CRISPR/Cas Systems

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In Vitro Directed Evolution of a Restriction Endonuclease with More Stringent Specificity
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In Vitro Directed Evolution of a Restriction Endonuclease with More Stringent Specificity

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Parallel High Throughput Single Molecule Kinetic Assay for Site-Specific DNA Cleavage
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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 生化学

背景情况:

  • 包括Cas12e核酶在内的V型CRISPR-Cas系统是用于DNA操纵和基因组编辑的多功能工具.
  • 现有的Cas12e变种,如DPbCas12e和PlmCas12e,在体外显示出不同的DNA裂变活动,突出显示了家族的多样性.

研究的目的:

  • 通过识别和分析新成员来全面描述Cas12e家族.
  • 了解Cas12e家族内的多种酶性质和裂解功效的结构基础.

主要方法:

  • 六个新的Cas12e成员的识别和描述.
  • 对CRISPR-locus架构,PAM偏好和体外dDNA裂变活动的分析.
  • 对Cas12e变体的结构比较,重点关注NTSB域及其在DNA解中的作用.

主要成果:

  • 六个以前未报告的Cas12e成员被确定具有不同的CRISPR-locus架构,PAM偏好和分离效率.
  • 与其他变体相比,plmCas12e在cis-cleavage中表现出优越的跨裂变活性和较低的盐敏感性.
  • 结构分析揭示了NTSB域对于高盐度的DNA解的重要性,其中一些变体使用带电循环.

结论:

  • 结构元素的不同进化,如NTSB域,驱动Cas12e家族内的核酶多样性.
  • 这些结构变异有助于Cas12e核酶适应不同的环境条件,并增强其作为基因组编辑工具的实用性.