2-Substituted dATP Derivatives as Building Blocks for Polymerase-Catalyzed Synthesis of DNA Modified in the Minor Groove

Abstract

2'-Deoxyadenosine triphosphate (dATP) derivatives bearing diverse substituents (Cl, NH₂ , CH₃ , vinyl, ethynyl, and phenyl) at position 2 were prepared and tested as substrates for DNA polymerases. The 2-phenyl-dATP was not a substrate for DNA polymerases, but the dATPs bearing smaller substituents were good substrates in primer-extension experiments, producing DNA substituted in the minor groove. The vinyl-modified DNA was applied in thiol-ene addition and the ethynyl-modified DNA was applied in a CuAAC click reaction to form DNA labelled with fluorescent dyes in the minor groove.

Keywords: DNA modification; DNA polymerase; bioconjugation; fluorescent labelling; nucleotides.