Covalent cross-linking of duplex DNA using 4-thio-2'-deoxyuridine as a readily modifiable platform for introduction of reactive functionality into oligonucleotides.

Abstract

Full details of the template-directed covalent cross-linking of duplex oligodeoxynucleotides are presented. 4-Thio-2'-deoxyuridine was incorporated synthetically into a 17mer oligodeoxynucleotide, and the thiocarbonyl group of the modified base was alkylated with a variety of alpha-bromoacetyl-derivatized diamines. Covalent cross-linking was initiated by annealing the electrophilic probe oligomers with their complementary sequences, where a dG base was targeted at the position complementary to the modified 4-thio-2'-deoxyuridine. The sequence selectivity of cross-link formation as a function of tether topology and rigidity was examined, and the thermal stability of the modified duplexes was measured by UV melting experiments.