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  • 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, 10 mM DTT)
      1mM ATP
      1 μg RNA
      10 % (v/v) DMSO
      1 μM [32P]pCp
      1 μl (20 units) T4 RNA Ligase
    2. Incubate overnight at 16°C