Removal of pyrophosphate from 5’ triphosphorylated RNA using RppH (NEB #M0356)

1.    Set up the reaction as shown below-
Component  Volume 
 5’ triphosphorylated RNA  Up to 2 µg
 NEBuffer 2
 2 µl
 RNA 5’ Pyrophosphohydrolase (RppH)  2 µl
 Nuclease-free water  to 20 µl

2.    Incubate at 37°C for 30 minutes.

3.    Stop the reaction by adding EDTA to a final concentration of 20mM. Alternatively, you can directly proceed to RNA clean up.
Note: The EDTA chelates Mg++ ions so if you intend to follow the RppH reaction with another enzymatic reaction that uses Mg++ without reaction clean up, avoid adding EDTA. We recommend purifying RNA between different enzymatic steps to ensure optimal performance of the enzymes.