Introducing structural diversity in oligonucleotides via photolabile, convertible c5-substituted nucleotides

Chemically synthesized oligonucleotides are functionalized at defined sites while in their protected form on solid-phase supports via the incorporation of nucleotides containing masked alkyl carboxylic acids or alkylamines. The reactive functional groups are selectively revealed upon 365 nm irradiation of the appropriate o-nitrobenzyl based protecting group. For the photolabile nucleotide that reveals alkylamines, a combination of analytical methods revealed that very high yields (≥94%) of the oligonucleotide containing a single reactive functional group were obtained by using photolysis conditions that do not damage the biopolymer. Either functionalized nucleotide is efficiently coupled to the corresponding reaction partner via PyBOP mediated amide bond formation. Isolated yields of the conjugated oligonucleotides are ≥90%. This method is compatible with introducing multiple modified sites sequentially and is amenable to the synthesis of libraries of oligonucleotides in a combinatorial fashion containing nonnative nucleotides while bound to their solid supports.