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Mapping Adenines, Guanines, And Pyrimidines In Rna, Helen Donis-Keller, Allan Maxam, Walter Gilbert

Helen Donis-Keller

The positions of adenines, guanines, and pyrimidines can be determined by partial nuclease digestion of a terminally labeled RNA molecule. In urea, at elevated temperatures, RNase T1 generates a pattern reflecting cleavage at guanines while RNase U2 cleaves only at adenine. A limited alkaline hydrolysis provides a continuum of fragments derived from breaks at every phosphodiester bond. The reaction products are electrophoretically fractionated by size in adjacent lanes of a polyacrylamide gel. An autoradiograph of the gel displays the sequence up to 100 nucleotides from the end of the molecule, although uracil cannot as yet be distinguished from cytosine. These …