This paper describes a method for determining the sequence of RNA by partial nuclease digestion and electrophoresis. The technique involves labeling RNA at one end with 32P, then using RNase T1 and RNase U2 to generate fragments that can be separated by size on a polyacrylamide gel. RNase T1 cleaves RNA at guanines, while RNase U2 cleaves at adenines. Limited alkaline hydrolysis produces fragments that can be compared with the nuclease digestion products to determine the positions of all nucleotides. The method was tested on yeast 5.8S ribosomal RNA, which is known to have a specific sequence. The results showed that the technique can determine the positions of adenines, guanines, and pyrimidines in RNA sequences, although uracil cannot be distinguished from cytosine. The method is useful for sequencing RNA and has been applied to other RNA molecules. The study also discusses the limitations of the technique, such as the inability to distinguish uracil from cytosine, and the need for further research to develop methods that can distinguish these two nucleotides. The work was supported by a grant from the National Institute of General Medical Sciences.This paper describes a method for determining the sequence of RNA by partial nuclease digestion and electrophoresis. The technique involves labeling RNA at one end with 32P, then using RNase T1 and RNase U2 to generate fragments that can be separated by size on a polyacrylamide gel. RNase T1 cleaves RNA at guanines, while RNase U2 cleaves at adenines. Limited alkaline hydrolysis produces fragments that can be compared with the nuclease digestion products to determine the positions of all nucleotides. The method was tested on yeast 5.8S ribosomal RNA, which is known to have a specific sequence. The results showed that the technique can determine the positions of adenines, guanines, and pyrimidines in RNA sequences, although uracil cannot be distinguished from cytosine. The method is useful for sequencing RNA and has been applied to other RNA molecules. The study also discusses the limitations of the technique, such as the inability to distinguish uracil from cytosine, and the need for further research to develop methods that can distinguish these two nucleotides. The work was supported by a grant from the National Institute of General Medical Sciences.