通过信号放大 ribozymes 来检测 MicroRNA 的特定序列
Jörg S Hartig1, Imke Grüne, S Hani Najafi-Shoushtari
1Kekulé Institut für Organische Chemie und Biochemie, Universität Bonn, Gerhard-Domagk-Strasse 1, 53121 Bonn, Germany.
Journal of the American Chemical Society
|January 22, 2004
概括
我们开发了一种新的毛 ribozyme 系统,用于敏感检测微RNA (小内源RNA分子). 这种方法为检测微小量的目标核酸提供了显著增强的灵敏度.
科学领域:
- 分子生物学分子生物学
- 生物化学 生物化学
- 核酸化学的核酸化学
背景情况:
- 微RNA (miRNA) 是小的内源性RNA分子,对基因调节至关重要.
- 精确检测miRNAs对于理解基因表达和疾病至关重要.
- 现有的检测方法,如分子信标,在灵敏度上有局限性.
研究的目的:
- 设计和描述一种用于特定序列核酸检测的新型毛核酶系统.
- 与传统方法相比,提高miRNA检测的灵敏度.
- 为了证明这个系统在复杂样本中检测低度miRNA的实用性.
主要方法:
- 由特定的外部寡核酸激发的针头核酶的设计.
- 利用 ribozyme 系统的内在信号放大特性.
- 该系统的应用用于检测特定的microRNAs.
- 在混合RNA样本中测试灵敏度和特异性.
主要成果:
- 设计的针头核糖酶表现出由外部寡核酸引起的序列特异性诱导.
- 该系统的灵敏度至少比标准分子信标增加了一个数量级.
- 在低度下,即使在混合其他miRNA序列中,也可以实现可靠和灵敏地检测相关的microRNAs.
结论:
- 开发的毛核酶系统提供了一种高度敏感和特定的方法来检测小核酸.
- 这项技术比现有的分子信标方法有了显著的改进.
- 这些探测器对需要检测微小数量的DNA或RNA,特别是miRNA的应用具有前景.
相关概念视频
MicroRNAs
MicroRNA (miRNA) are short, regulatory RNA transcribed from introns—non-coding regions of a gene—or intergenic regions—stretches of DNA present between genes. Several processing steps are required to form biologically active, mature miRNA. The initial transcript, called primary miRNA (pri-mRNA), base-pairs with itself forming a stem-loop structure. Within the nucleus, an endonuclease enzyme, called Drosha, shortens the stem-loop structure into hairpin-shaped pre-miRNA. After the pre-miRNA ends...
Riboswitches
Riboswitches are non-coding mRNA domains that regulate the transcription and translation of downstream genes without the help of proteins. Riboswitches bind directly to a metabolite and can form unique stem-loop or hairpin structures in response to the amount of the metabolite present. They have two distinct regions – a metabolite-binding aptamer and an expression platform.
The aptamer has high specificity for a particular metabolite which allows riboswitches to specifically regulate...
The aptamer has high specificity for a particular metabolite which allows riboswitches to specifically regulate...
DNA Microarrays
Microarrays are high-throughput and relatively inexpensive assays that can be automated to analyze large quantities of data at a time. They are used in genome-wide studies to compare gene or protein expression under two varied conditions, such as healthy and diseased states. Microarrays consist of glass or silica slides on which probe molecules are covalently attached through surface functionalization. Most commonly, the slides are prepared through the chemisorption of silanes to silica...
RNA-seq
RNA sequencing, or RNA-Seq, is a high-throughput sequencing technology used to study the transcriptome of a cell. Transcriptomics helps to interpret the functional elements of a genome and identify the molecular constituents of an organism. Additionally, it also helps in understanding the development of an organism and the occurrence of diseases.
Before the discovery of RNA-seq, microarray-based methods and Sanger sequencing were used for transcriptome analysis. However, while microarray-based...
Before the discovery of RNA-seq, microarray-based methods and Sanger sequencing were used for transcriptome analysis. However, while microarray-based...
Ribosome Profiling
Ribosome profiling or ribo-sequencing is a deep sequencing technique that produces a snapshot of active translation in a cell. It selectively sequences the mRNAs protected by ribosomes to get an insight into a cell’s translation landscape at any given point in time.
Applications of ribosome profiling
Ribosome profiling has many applications, including in vivo monitoring of translation inside a particular organ or tissue type and quantifying new protein synthesis levels.
The technique helps...
Applications of ribosome profiling
Ribosome profiling has many applications, including in vivo monitoring of translation inside a particular organ or tissue type and quantifying new protein synthesis levels.
The technique helps...
MicroRNAs
MicroRNA (miRNA) are short, regulatory RNA transcribed from introns (non-coding regions of a gene) or intergenic regions (stretches of DNA present between genes). Several processing steps are required to form biologically active, mature miRNA. The initial transcript, called primary miRNA (pri-mRNA), base-pairs with itself, forming a stem-loop structure. Within the nucleus, an endonuclease enzyme, called Drosha, shortens the stem-loop structure into hairpin-shaped pre-miRNA. After the pre-miRNA...


