title: Synthesis of building blocks and oligonucleotides with {T}N3-alkylene-N3{T} cross-links.
abstract: This unit describes two methods to directly prepare oligonucleotide duplexes containing an N3thymidine-alkylene-N3thymidine inter-strand cross-link. The inter-strand cross-link can be engineered into the duplex with a number of possible orientations. Both methods require the preparation of a protected thymidine dimer where the N3 atoms of the two nucleosides are covalently attached by an alkyl linker. This linker is prepared starting from a protected diol using two successive alkylation reactions under basic conditions to accomplish the alkylation selectively at the N3 atom of the nucleoside. The chain length of the cross-link can be varied based on the selection of the diol used in the dimer synthesis. The solid-phase mono-phosphoramidite approach involves oligonucleotide synthesis with 3'-O-phosphoramidites, on-column removal of a 3'-O-tert-butyldimethylsilyl protecting group, and continued oligonucleotide synthesis with 5'-O-phosphoramidites. The bis-phosphoramidite approach does not require synthesis with 5'-O-phosphoramidites. At the end of synthesis using either method, the N3thymidine-alkylene-N3thymidine inter-strand cross-linked oligonucleotides can be removed from the solid-support and purified using standard techniques (ion-exchange HPLC) in yields sufficient for various structural studies and repair assays.
authors.affiliation: Department of Chemistry and Biochemistry, Concordia University, Montreal, Quebec, Canada.
authors.forename: Gang
authors.initials: G
authors.lastname: Sun
authors.forename: Anne M
authors.initials: AM
authors.lastname: Noronha
authors.forename: Paul S
authors.initials: PS
authors.lastname: Miller
authors.forename: Christopher J
authors.initials: CJ
authors.lastname: Wilds
mesh_terms: D000477:Alkylating Agents / Q000737:chemistry; D002850:Chromatography, Gel / Q000379:methods; D004247:DNA / Q000737:chemistry; D004260:DNA Repair; D009841:Oligonucleotides / Q000138:chemical synthesis* / Q000737:chemistry; D009943:Organophosphorus Compounds / Q000138:chemical synthesis* / Q000737:chemistry; D060327:Solid-Phase Synthesis Techniques / Q000379:methods; D013936:Thymidine / Q000031:analogs & derivatives* / Q000737:chemistry
chemical_list: D000477:Alkylating Agents; D009841:Oligonucleotides; D009943:Organophosphorus Compounds; C434331:phosphoramidite; D004247:DNA; D013936:Thymidine