The Enzymic Addition of Poly(A) to the 3′‐End of RNA using Bacteriophage MS 2 RNA as a Model System

Abstract

ATP : RNA adenyltransferase, purified from Escherichia coli, was used to add a series of adenosine residues to the 3'-end of MS2RNA. Incubations of the order of a few minutes at 37 degrees C were sufficient for synthesis of a short poly(A) chain that did not appreciably alter the hydrodynamic or electrophoretic properties of MS2 RNA. The size of the poly(A) tails was estimated by gel electrophoresis after prior hydrolysis of the primer RNA with pancreatic ribonuclease. These results were in good agreement with the values calculated on the basis of the relative amount of incorporated AMP. After the addition of a short poly(A) tail, approximately 50% of the treated material binds specifically to an oligo(dT)-cellulose column. The majority of the recovered poly(a)-containing RNA was still intact, as shown by analysis on polyacrylamide gel. After incubations beyond 6 min, slowly sedimenting material, also showing reduced electrophoretic mobility, was formed. Presumably this material corresponds to RNA chains to which long poly(A) tails are linked.