3 dutp

ATP, UTP, CTP, GTP, 3′-dUTP, 3′-dATP, 3′-dGTP, 3′-dCTP, 2′-dATP, 2′-dCTP, 2′-dGTP, 2′-dTTP, 2′-F-CTP, 2′-F-GTP, 2′-NH₂-GTP, 2′-NH₂-CTP, 2′-F-ATP, 2′-F-UTP, ara-GTP, ara-ATP, 7-deaza-ATP, ara-CTP, ara-UTP, 2′-azido-GTP, and 2′-azido-CTP were purchased as 100 mM solutions from TriLink Biotechnologies (San Diego, CA). 2′-C-ethynyl-7-deaza-ATP was purchased from Carbosynth (San Diego, CA). Urea, taurine, dithiothreitol (DTT), imidazole, MgCl₂, and IPTG (isopropyl-β-d-thiogalactopyranoside) were purchased from Sigma (St. Louis, MO). LB medium, bovine serum albumin (BSA), NaCl, Triton X-100, glycerol, protease inhibitor cocktail, Tris-HCl (pH 7.5), and HisPur Ni-NTA agarose resin were purchased from Thermo Fisher Scientific (Waltham, MA). An In-Fusion HD cloning kit and Stellar competent cells were purchased from Clontech (Mountain View, CA). The pMal-c5X vector was purchased from New England Bioscience (NEB; Ipswich, MA). Fluorescently labeled RNA (Cy5.5-RNA) and unlabeled DNA oligonucleotides were chemically synthesized and purified by high-performance liquid chromatography by Integrated DNA Technologies (Coralville, IA). 2′-C-Me-ATP, 2′-C-Me-GTP, 2′-F-2′-C-Me-GTP, 4′-azido-GTP, 2′-F-2′-C-Me-CTP, 2′-C-Me-CTP, 4′-azido-CTP, 2′-F-2′-C-Me-UTP, 2′-C-Me-UTP, 2′-C-ethynyl-UTP, and 4′-azido-UTP were custom synthesized in-house.