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関連する概念動画

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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プログラムされた自己分裂DNAzymeを使用して増幅された核酸センサー.

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
まとめ

新しいターゲットアシストドセルフ・クリバージ (TASC) 探査機は,同熱で酵素フリーなDNA/RNA検出を可能にします. この触媒反応は,標的配列情報を増幅し,光を用いた単一核酸差別を可能にします.

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科学分野:

  • 分子生物学は分子生物学である.
  • バイオテクノロジー バイオテクノロジー
  • 核酸化学 核酸化学について

背景:

  • 現在の核酸検出方法は,PCR増幅を含む複雑なプロトコルと複数の反応剤を必要とします.
  • DNAやRNAの標的を検出するよりシンプルで,より敏感で,より特殊な検出方法が必要である.
  • 触媒性核酸探査機は,信号増幅と簡素化された検出スキームの可能性を秘めています.

研究 の 目的:

  • ターゲットアシストドセルフクリバージ (TASC) による核酸検出のための新しい探査システムの設計と特徴付け.
  • ターゲットシーケンス情報の同熱,酵素フリー増幅のためのTASCプローブの能力を実証する.
  • ターゲット配列における単核酸差異の敏感な差別化のための光報告型TASC探査機を開発する.

主な方法:

  • 標的結合部位とDNA酵素ドメインを併用したTASCプローブの設計.
  • 標的DNA/RNAが触媒として作用するTASC反応機構の調査.
  • 断裂部位を横断するフォースター共振エネルギー転送 (FRET) ペア (フローレスセイン/ダブシル) を利用した光報告型TASCプローブの開発.

主要な成果:

  • TASC探査機は,標的DNA/RNAとハイブリッド化すると,効率的な自己分裂をします.
  • 自己分裂反応は触媒的であり,探査機は基板として,標的は触媒として作用し,製品の放出につながります.
  • 光を報告するTASC探査機は,混ぜて読み解く検出と,同熱,酵素/反応剤のない条件下で,ターゲット配列の単核酸変異の差別化を可能にします.

結論:

  • TASC探査機は,核酸検出と配列情報増幅のための新しい効率的なプラットフォームを表しています.
  • TASCシステムはPCR以外の単純な同熱条件下で動作し,酵素や反応剤の必要性を排除します.
  • 光報告型TASC探査機は,単核酸多形態化 (SNP) 分析を含む,核酸標的の敏感で特異的な検出のための有望なアプローチを提供します.