费斯特共振能量转移
适体
生物传感器
核酶
变构调节
化学
核糖开关
配体(生物化学)
核糖核酸
分析物
组合化学
荧光
生物物理学
纳米技术
生物化学
材料科学
生物
受体
分子生物学
非编码RNA
物理
物理化学
基因
量子力学
作者
David Rueda,Nils G. Walter
出处
期刊:Humana Press eBooks
[Humana Press]
日期:2006-06-24
卷期号:: 289-310
被引量:35
标识
DOI:10.1385/1-59745-069-3:289
摘要
Biosensors are devices that amplify signals generated from the specific interaction between a receptor and an analyte of interest. RNA structural motifs called aptamers have recently been discovered as receptor components for biosensors owing to the ease with which they can be evolved in vitro to bind a variety of ligands with high specificity and affinity. By coupling an aptamer as allosteric control element to a catalytic RNA such as the hammerhead ribozyme, ligand binding is transduced into a catalytic event. We have made use of fluorescence resonance energy transfer (FRET) to further amplify ligand induced catalysis into an easily detectable fluorescence signal. This chapter reviews in detail the methods and protocols to prepare a theophylline specific aptazyme and to label its substrate with fluorophores. We also include detailed protocols to characterize by FRET the binding affinity of the target, theophylline, as well as the external substrate to the aptazyme. The chapter should therefore facilitate the implementation of RNA-based biosensor components for other analytes of interest.
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