Enzymatic Joining of Polynucleotides

Abstract

A new method for the assay of polynucleotide joining activity is described; it measures the conversion of 3H-labeled d(A-T) copolymer with 3'-hydroxyl and 5'-phosphoryl termini to a form resistant to exonuclease III. The method is rapid and precise and is suitable for assay of crude cell extracts. The product of the action of joining enzyme on the d(A-T) copolymer has the properties of a circular molecule. The optimal chain length of the d(A-T)n substrate is approximately 1000 nucleotides.