Sequence analysis of T1 ribonuclease fragments of 18S ribosomal RNA by 5′-terminal labeling, partial digestion, and homochromatography fingerprinting

The method employed to determine the sequence of a T1 RNase fragment, A-A-A-A-A-U-A-A-C-A-A-U-A-C-A-Gp, from Novikoff rat hepatoma 18S ribosomal RNA is described. This method is applicable to any oligoribonucleotide produced by specific endonucleases that leave the newly cleaved 5'-end free for labeling with polynucleotide kinase and gamma-(32p)-ATP. The (32p)-labeled oligoribonucleotide is subjected to partial endonucleolytic digestion and fractionated by two-dimensional homochromatography fingerprinting. The nucleotide sequence is determined by following mobility shifts of the labeled and partially digested oligoribonucleotides in homochromatography fingerprinting.