title: Strain-promoted "click" chemistry for terminal labeling of DNA.
abstract: 1,3-Dipolar [3 + 2] cycloaddition between azides and alkynes--an archetypal "click" chemistry--has been used increasingly for the functionalization of nucleic acids. Copper(I)-catalyzed 1,3-dipolar cycloaddition reactions between alkyne-tagged DNA molecules and azides work well, but they require optimization of multiple reagents, and Cu ions are known to mediate DNA cleavage. For many applications, it would be preferable to eliminate the Cu(I) catalyst from these reactions. Here, we describe the solid-phase synthesis and characterization of 5'-dibenzocyclooctyne (DIBO)-modified oligonucleotides, using a new DIBO phosphoramidite, which react with azides via copper-free, strain-promoted alkyne-azide cycloaddition (SPAAC). We found that the DIBO group not only survived the standard acidic and oxidative reactions of solid-phase oligonucleotide synthesis (SPOS), but that it also survived the thermal cycling and standard conditions of the polymerase chain reaction (PCR). As a result, PCR with DIBO-modified primers yielded "clickable" amplicons that could be tagged with azide-modified fluorophores or immobilized on azide-modified surfaces. Given its simplicity, SPAAC on DNA could streamline the bioconjugate chemistry of nucleic acids in a number of modern biotechnologies.
authors.affiliation: Department of Chemistry, University of Minnesota, 207 Pleasant Street SE, Minneapolis, Minnesota 55455, United States.
authors.forename: Isaac S
authors.initials: IS
authors.lastname: Marks
authors.forename: Jun Sung
authors.initials: JS
authors.lastname: Kang
authors.forename: Brady T
authors.initials: BT
authors.lastname: Jones
authors.forename: Kevin J
authors.initials: KJ
authors.lastname: Landmark
authors.forename: Andrew J
authors.initials: AJ
authors.lastname: Cleland
authors.forename: T Andrew
authors.initials: TA
authors.lastname: Taton
mesh_terms: D000480:Alkynes / Q000737:chemistry*; D001386:Azides / Q000737:chemistry*; D057930:Click Chemistry / Q000379:methods*; D003500:Cyclization; D004247:DNA / Q000138:chemical synthesis / Q000737:chemistry*; D005456:Fluorescent Dyes / Q000737:chemistry; D009841:Oligonucleotides / Q000138:chemical synthesis / Q000737:chemistry*; D016133:Polymerase Chain Reaction
chemical_list: D000480:Alkynes; D001386:Azides; D005456:Fluorescent Dyes; D009841:Oligonucleotides; C042001:1-octyne; D004247:DNA