Lincoln Scott - RNA structure determination by NMR.

Document created by Lincoln Scott on Aug 22, 2014
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  Methods Mol   Biol. 2008;452:29-61. doi:   10.1007/978-1-60327-159-2_2.


  This chapter reviews the methodologies for RNA structure   determination by liquid-state nuclear magnetic resonance (NMR).   The routine production of milligram quantities of isotopically   labeled RNA remains critical to the success of NMR-based   structure studies. The standard method for the preparation of   isotopically labeled RNA for structural studies in solution is in   vitro transcription from DNA oligonucleotide templates using T7   RNA polymerase and unlabeled or isotopically labeled nucleotide   triphosphates (NTPs). The purification of the desired RNA can be   performed by either denaturing polyacrylamide gel electrophoresis   (PAGE) or anion-exchange chromatography. Our basic strategy for   studying RNA in solution by NMR is outlined. The topics covered   include RNA resonance assignment, restraint collection, and the   structure calculation process. Selected examples of NMR spectra   are given for a correctly folded 30 nucleotide-containing RNA.

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