Triazole linking for preparation of a next-generation sequencing library from single-stranded DNA

Abstract

Next-generation sequencing of single-stranded DNA (ssDNA) is attracting increased attention from a wide variety of research fields. Accordingly, various methods are actively being tested for the efficient adaptortagging of ssDNA. We conceived a novel chemoenzymatic method termed terminal deoxynucleotidyl transferase (TdT)-assisted, copper-catalyzed azidealkyne cycloaddition (CuAAC)-mediated ssDNA ligation (TCS ligation). In this method, TdT is used to incorporate a single 3'-azide-modified dideoxyribonucleotide onto the 3'-end of target ssDNA, followed by CuAAC-mediated click ligation of the azideincorporated 3'-end to a 5'-ethynylated synthetic adaptor. This report presents the first proof-ofprinciple application of TCS ligation with its use in the preparation of a next-generation sequencing library.