The isolation of 3′-terminal polynucleotides from RNA molecules

Abstract

The 3′-terminal polynucleotides obtained by nuclease digestion of large RNA's can be selectively bound to columns of cellulose derivatives containing covalently bound dihydroxyboryl groups. The binding results from complex formation between the dihydroxyboryl groups and the 2′,3′-diol moieties of the terminal polynucleotides, and is effected at pH 8–9 in the presence of Mg 2+. The rest of the polynucleotide fragments in the digest are not bound under these conditions and, after their removal by elution, the terminal polynucleotides are recovered by changing the pH of the eluting solvent to 5–6. The method has been applied to the isolation of the terminal polynucleotides obtained by ribonuclease T 1 digestion of the RNA's of bacteriophages f2, Qβ, and GA as well as rRNA and tRNA.