Protocol for conversion of Uridine to Pseudouridine using Sce PUS1 (NEB #M0526)

1. Prepare 100 µl reaction as described below:
   
   

   Components	100 µl Reaction
   RNA	up to 1 µg
   NEBuffer r1.1 (10X)	10 µl
   Sce PUS1	1 µl (100 pmoles)
   Nuclease-free water	to 100 µl

   
   
   
2. Incubate at 30°C for 2 hours
   
   
3. The reaction may be stopped by the addition of 1 µl of NEB Proteinase K and 15 min incubation at 37°C. Subsequent RNA purification using the appropriate NEB Monarch RNA Cleanup Kit can be used to clean up the RNA for downstream applications.