通过体外转录使用扩展遗传密码子制备位点特异性自旋标记 RNA。
Preparation of Site-Specifically Spin-Labeled RNA by in Vitro Transcription Using an Expanded Genetic Alphabet.
机构信息
Department of Chemistry, University of Cologne, Köln, Germany.
出版信息
Methods Mol Biol. 2022;2439:223-240. doi: 10.1007/978-1-0716-2047-2_15.
Recent advances in pulsed electron paramagnetic resonance (EPR) spectroscopy enable studying structure and folding of nucleic acids. An efficient introduction of spin labels at specific positions within the oligonucleotide sequence is a prerequisite. We here present a step-by-step guide to synthesize long RNA oligonucleotides bearing spin labels at specific positions within the sequence. RNA preparation is achieved enzymatically via in vitro transcription using an expanded genetic alphabet. Highly structured, several hundred nucleotides long RNAs with two nitroxide spin labels at specific positions can be prepared by this method.
近年来，脉冲电子顺磁共振（EPR）光谱技术的发展使得研究核酸的结构和折叠成为可能。在寡核苷酸序列的特定位置引入自旋标记物是前提条件。本研究提供了一种在寡核苷酸序列的特定位置引入自旋标记物的长 RNA 寡核苷酸的分步合成指南。通过使用扩展遗传密码的体外转录酶法实现 RNA 的制备。通过这种方法可以制备具有两个特定位置的硝酮自旋标记的高度结构化的几百个核苷酸长的 RNA。
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