Use of specific endonuclease cleavage in RNA sequencing - an enzymic method for distinguishing between cytidine and uridine residues

摘要

The extracellular ribonuclease I of the common slime mold Physarum polycephalum1 (RNase Phy₁), which has recently been purified to homogeneity2, has been used to distinguish between C and U residues in 3'-end-labeled oligoribonucleotides. As shown by Bargetzi and coworkers2, this enzyme exhibits strong cleavage preference for U-N over C-N and N-C over N-U bonds. In the present paper, conditions are being detailed, which enable one to deduce the sequences of rather large, pyrimidine-rich, terminally labeled oligonucleotides by partial digestion with RNases U₂, A, and Phy₁, followed by resolution of the cleavage products by size. The techniques described in this and a previous3 communication provide a direct means for identifying A, G, C, and U residues in end-labeled polyribonucleotides.