title: Fluorescently labeling synthetic RNAs.
abstract: This protocol covers the steps required to incorporate N-hydroxysuccinamide (NHS) functionalized fluorophores into synthetic RNAs containing a residue derivatized with a primary amine. This method has been widely used to label RNA oligonucleotides that are used directly, targeted to a complementary RNA using base pairing rules, or covalently ligated to a RNA of interest (Ha et al., 1999; Hodak et al., 2005; Baum and Silverman, 2007; Sattint et al., 2008; Akiyama and Stone, 2009; Solomatin and Herschlag, 2009). While this technique is quite general, the details of a particular experiment can vary, therefore, it is always important to keep in mind that other labeling strategies are available and should potentially be considered.
authors.affiliation: Department of Chemical Engineering, Stanford University, Stanford, CA, USA; Department of Biochemistry, Stanford University, Stanford, CA, USA.
authors.forename: Max
authors.initials: M
authors.lastname: Greenfeld
authors.forename: Daniel
authors.initials: D
authors.lastname: Herschlag
mesh_terms: D000577:Amides / Q000737:chemistry; D011232:Chemical Precipitation; D031541:Fluorescence Resonance Energy Transfer / Q000379:methods*; D005456:Fluorescent Dyes / Q000737:chemistry*; D009841:Oligonucleotides / Q000737:chemistry; D012313:RNA / Q000737:chemistry*; D013194:Staining and Labeling / Q000379:methods; D013386:Succinates / Q000737:chemistry
chemical_list: D000577:Amides; D005456:Fluorescent Dyes; D009841:Oligonucleotides; D013386:Succinates; D012313:RNA; C011116:succinamide
keywords: Fluorescence resonance energy transfer (FRET); Gel purification; Large-scale labeling reaction; NHS-activated fluorophores; Precipitate unlabeled oligonucleotide; Synthetic RNAs