XNAzyme-mediated RNA Cleavage Analysis

1 uM “Sub_Ebo” RNA was reacted with XNAzyme “FR6_1” under high magnesium selection conditions (20 h, 17 °C) and the 5’ RNA cleavage product purified by Urea-PAGE, extracted from macerated gel using a 0.22 uM centrifugal filter (Corning, USA) as described previously^{7 (link)} and ethanol precipitated. The cleavage product was resuspended in nuclease-free water and analysed by MALDI-ToF mass spectrometry using an Ultraflex III TOF-TOF instrument (Bruker Daltonik, Germany) in positive ion mode as described previously^{2 (link)}. Expected masses were calculated using the Mongo Oligo Mass Calculator v.2.06. The 5’ “6FAM” (6-Carboxyfluorescien)(IDT, USA) has a mass of 537.5 Da.