非自然な塩基対の効率的に複製され,転写されたクラスを使用して,DNAとRNAのサイト固有のラベリングを行います
Young Jun Seo1, Denis A Malyshev, Thomas Lavergne
1Department of Chemistry and Center for Protein and Nucleic Acid Research, The Scripps Research Institute, 10550 N. Torrey Pines Road, La Jolla, California 92037, USA.
Journal of the American Chemical Society
|October 11, 2011
まとめ
研究者らは,サイト固有のDNAとRNAのラベル付けのための新しい非自然な塩基対を開発しました. これらの遺伝子組み換え核酸は,効率的な酵素合成と合成後の機能化を可能にし,さまざまな用途に使用されます.
科学分野:
- バイオケミストリー バイオケミストリー
- 分子生物学は分子生物学である.
- 合成生物学 合成生物学とは
背景:
- 核酸のサイト固有のラベル付けは,生体物理学,核酸進化,ナノ材料,バイオセンサの応用に不可欠です.
- 不自然な塩基対で遺伝子アルファベットを拡張することは,新しいラベリング戦略を容易にします.
研究 の 目的:
- サイト固有のDNAとRNAの改変のためのプロパルギラミンリンカーを用いて,改変された非自然な塩基対 (d) 5SICSと (d) MMO2を合成および分析する.
- バックボーンチオレーションと増幅後のラベル付けのためのα-フォスフォロチオ酸エート変種を評価する.
- 酵素合成におけるこれらの改変核酸の効率と忠誠度を評価する.
主な方法:
- (d) 5SICSと (d) MMO2のリボ変種とデオキシリボ変種を,機能的リンク子で合成する.
- 安定状態運動学とポリメラーゼ連鎖反応 (PCR) を用いたデオキシヌクレオチドの特徴化.
- モデルRNAとtRNAの転写によるリボヌクレオシドの特徴化.
主要な成果:
- 改変された (d) 5SICSと (d) MMO2核酸の合成と特徴づけに成功した.
- 酵素合成中のまたは後のDNAとRNAの効率的でサイト固有のラベル付けが実証されています.
- リンカー結合に対するニュクレオチドおよびポリメラーゼ固有の感受性が特定されましたが,全体的に高い効率と忠誠性が達成されました.
結論:
- 開発された非自然な塩基対 (d) 5SICSと (d) MMO2は,サイト特異的に改変されたDNAとRNAを生成するのに適しています.
- これらの改変核酸は,多種多様なグループで機能化され,実用的な応用が可能である.
- これらの戦略は,高度な研究とバイオテクノロジーのための核酸のラベル付けに多岐に渡る経路を提供している.
関連する概念動画
Labeling DNA Probes
DNA probes are fragments of DNA labeled with a reporter tag to enable their detection or purification. The resulting labeled DNA probes can then hybridize to target nucleic acid sequences through complementary base-pairing, and may be used to recover or identify these regions.
Radioisotopes, fluorophores, or small molecule binding partners like biotin or digoxigenin, are the most widely used reporter tags for labeling DNA probes. These labels can be attached to the probe DNA molecule via...
Radioisotopes, fluorophores, or small molecule binding partners like biotin or digoxigenin, are the most widely used reporter tags for labeling DNA probes. These labels can be attached to the probe DNA molecule via...
In-situ Hybridization
In situ hybridization (ISH) is a technique used to detect and localize specific DNA or RNA molecules in cells, tissue, or tissue sections using a labeled probe. The technique was first used in 1969 for the investigation of nucleic acids. It is currently an essential tool in scientific research and clinical settings, especially for diagnostic purposes.
Types of probes and labels
A probe is a complementary strand of DNA or RNA that binds to corresponding nucleotide sequences in a cell. Many...
Types of probes and labels
A probe is a complementary strand of DNA or RNA that binds to corresponding nucleotide sequences in a cell. Many...
Conservative Site-specific Recombination and Phase Variation
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.
The recognition sites for Cre recombinase called LoxP...
The recognition sites for Cre recombinase called LoxP...
Sanger Sequencing
DNA sequencing is a fundamental technique that is routinely used in the biological sciences. This method can be applied to a range of questions at different scales - from the sequencing of a cloned DNA fragment or the study of a mutation in a gene up to whole-genome sequencing. However, despite the widespread use of sequencing today, it was not until 1977 that Fredrick Sanger and his collaborators developed the chain-termination method to decode DNA sequences. It relies on the separation of a...
Maxam-Gilbert Sequencing
In the same year as the discovery of the Sanger sequencing method, another group of scientists, Allan Maxam and Walter Gilbert, demonstrated their chemical-cleavage method for DNA sequencing. The Maxam-Gilbert method relies on using different chemicals that can cleave the DNA sequence at specific sites, the separation of resulting DNA fragments of variable size using electrophoresis, and deciphering the DNA sequence from the resulting gel bands.
Challenges of the Maxam-Gilbert Method
The...
Challenges of the Maxam-Gilbert Method
The...
Next-generation Sequencing
The first human genome sequencing project cost $2.7 billion and was declared complete in 2003, after 15 years of international cooperation and collaboration between several research teams and funding agencies. Today, with the advent of next-generation sequencing technologies, the cost and time of sequencing a human genome have dropped over 100 fold.
Next-Generation Sequencing Methods
Although all next-generation methods use different technologies, they all share a set of standard features.
Next-Generation Sequencing Methods
Although all next-generation methods use different technologies, they all share a set of standard features.


