3´ End Labeling of RNA using T4 RNA Ligase 1

Introduction

Set up the labeling reaction at room temperature until the ligase is added to avoid DMSO precipitation. The addition of DMSO is essential.

Protocol

1. Set up a 30 μl reaction as follows:
   1 X Reaction Buffer (50 mM Tris-HCl, pH 7.8, 10 mM MgCl₂, 1 mM DTT)
   1mM ATP
   1 μg RNA
   10 % (v/v) DMSO
   1 μM [³²P]pCp
   1 μl (10 units) T4 RNA Ligase
2. Incubate overnight at 16°C