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