Use of specific endonuclease cleavage in RNA sequencing-an enzymic method for distinguishing between cytidine and uridine residues.
The extracellular ribonuclease I of the common slime mold physarum polycephalum (RNase Phy1), which has recently been purified to homogeneity, has been used to distinguish between C and U residues in 3'-end-labeled oligoribonucleotides. As shown by Bargetzi and coworkers, 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 previous communication provide a direct means for identifying A, G, C, and U residues in end-labeled polyribonucleotides.
D001483:Base Sequence; D003562:Cytidine / Q000032:analysis*; D004720:Endonucleases / Q000378:metabolism; D008722:Methods; D008970:Molecular Weight; D010804:Physarum / Q000201:enzymology; D012313:RNA; D012260:Ribonucleases / Q000378:metabolism; D013379:Substrate Specificity; D014529:Uridine / Q000032:analysis*
D003562:Cytidine; D012313:RNA; D004720:Endonucleases; D012260:Ribonucleases; D014529:Uridine