Long-range scalar (5)J(H1',F) couplings were observed in 5-fluoropyrimidine-substituted RNA. We developed a novel (SE)-E-3-F-19-alpha,beta-edited NOESY experiment for quantitation of these long-range scalar (5)J(H1',F) couplings, where the J-couplings can be extracted from inspection of intraresidual (H1',H6) NOE cross-peaks. Quantum chemical calculations were exploited to investigate the relation between scalar couplings and conformations around the glycosidic bond in oligonucleotides. The theoretical dependence of the observed (5)J(H1',F) couplings on the torsion angle chi can be described by a generalized Karplus relationship. The corresponding density functional theory (DFT) analysis is outlined. Additional NMR experiments facilitating the resonance assignments of 5-fluoropyrimidine-substituted RNAs are described, and chemical shift changes due to altered shielding in the presence of fluorine-19 (F-19) are presented.