FAQ: How much RNase 4 (NEB #M1284) should I use to digest RNA for LC-MS/MS nucleotide sequence mapping?

The addition of 1 µl of RNase 4 (NEB #M1284) (50,000 U/ml) yields maximal sequence coverage for 10 µg of unmodified IVT RNAs 1-5 kb in length in a 30 µl reaction (1X NEBuffer r1.1, 1 M Urea) incubated at 37°C for 1 h. However, similar to using RNase T1 or bovine pancreatic RNase A, the ideal number of RNase 4 units needed can be higher or lower based on the target RNA. RNase 4 can be diluted in 1X NEBuffer r1.1, included as a 10X solution.

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RNase 4