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相关概念视频

PCR01:32

PCR

Overview
Long-patch Base Excision Repair01:02

Long-patch Base Excision Repair

Since the discovery of the two BER pathways, there has been a debate about how a cell chooses one pathway over the other and the factors determining this selection. Numerous in vitro experiments have pointed out multiple determinants for the sub-pathway selection. These are:
Fixing Double-strand Breaks02:04

Fixing Double-strand Breaks

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...
PCR - Polymerase Chain Reaction01:32

PCR - Polymerase Chain Reaction

Overview
Restriction Enzymes01:11

Restriction Enzymes

Restriction enzymes are bacterial enzymes used to cut DNA in a sequence-specific manner. To cleave DNA, they bind to specific palindromic sequences called restriction sites. Such palindromic DNA sequences or inverted repeats are commonly found in regions of functional significance, such as the origin of replication, gene operator sites, and regions containing transcription termination signals.
The host bacteria protect their own genomic DNA from these enzymes by methylating these sites. Some...
Proofreading01:31

Proofreading

Synthesis of new DNA molecules is carried out by the enzyme DNA polymerase, which adds nucleotides on the daughter strand complementary to the template DNA strand. DNA polymerase has a higher affinity to add the correct base and ensures fidelity during DNA replication. Furthermore,  it exhibits proofreading activity during replication, using an exonuclease domain that cuts off incorrect nucleotides from the nascent DNA strand.
Errors During Replication are Corrected by the DNA Polymerase Enzyme

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相关实验视频

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In Vitro Directed Evolution of a Restriction Endonuclease with More Stringent Specificity
09:16

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

Published on: March 25, 2020

扩大核酸感应使用编程自裂 DNAzyme 酶.

Shinsuke Sando1, Toshinori Sasaki, Keiichiro Kanatani

  • 1Department of Synthetic Chemistry and Biological Chemistry, Graduate School of Engineering, Kyoto University, Katsura, Nishikyo-ku, Kyoto 615-8510, Japan. ssando@sbchem.kyoto-u.ac.jp

Journal of the American Chemical Society
|December 18, 2003
PubMed
概括

一种新型的目标辅助自切割 (TASC) 探头可以实现异热,无酶的DNA/RNA检测. 这种催化反应放大了目标序列信息,并允许使用光进行单核酸区分.

科学领域:

  • 分子生物学分子生物学
  • 生物技术是生物技术.
  • 核酸化学的核酸化学

背景情况:

  • 目前的核酸检测方法通常需要复杂的协议,包括PCR放大和多种试剂.
  • 需要更简单,更敏感和更具体的检测方法来检测DNA和RNA标.
  • 催化核酸探针为信号放大和简化检测方案提供了潜力.

研究的目的:

  • 设计和表征一种基于目标辅助自分裂 (TASC) 的核酸检测新型探针系统.
  • 为了证明TASC探针对目标序列信息的同热,无酶放大功能的能力.
  • 开发一种光报告TASC探针,用于对目标序列中的单核酸差异进行敏感的歧视.

主要方法:

  • 一个TASC探针的设计,包括一个目标结合部位和一个DNA酶域.
  • 研究TASC反应机制,其中目标DNA/RNA作为催化剂.
  • 开发一种光报告TASC探测器,使用Förster共振能量转移 (FRET) 对 (光素/dabsyl) 穿过裂变部位.

主要成果:

  • 在与目标DNA/RNA杂交后,TASC探头经历了高效的自我分裂.
  • 自分裂反应是催化反应,探头作为基质,目标作为催化剂,导致产品释放.
  • 光报告TASC探测器可以在异热,无酶/无试剂条件下混合读取检测和区分目标序列中的单核酸变异.

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结论:

  • TASC探测器代表了用于核酸检测和序列信息放大的新高效平台.
  • 该TASC系统在简单的,非PCR,同热条件下运行,不需要酶和试剂.
  • 光报告TASC探针为敏感和特定检测核酸标提供了一个有希望的方法,包括单核酸多态 (SNP) 分析.