Versatile linker chemistry for synthesis of 3'-modified DNA

A general method is described for the solid phase supported synthesis of DNA containing 3'-terminal phosphodiesters modified with linkers bearing either amino, thiol, or hydroxyl groups. These products are all made from a common intermediate, obtained by the reaction of trimellitic anhydride chloride with aminopropyl CPG. The anhydride-derivatized support was then reacted with three appropriate bifunctional spacers, giving DMT-protected hydroxyl solid supports bearing the masked functionality as an ester, amide, or thioester. DNA synthesis was then performed, followed by ammonia cleavage and deprotection, giving the hydroxyl-, amino-, or thiol-functionalized DNA 3'-phosphate diesters, respectively. Test mononucleotides synthesized with each of the new supports were identical with control mononucleotides made with 5'-immobilized nucleosides and alkylhydroxyl, alkylamino, and alkylthio phosphoramidites. The new supports were then used to prepare several 3'-modified oligonucleotides, which were characterized by gel electrophoresis, HPLC, and MALDI mass spectroscopy. The amino- and thiol-functionalized DNAs were conjugated with chromophores, and purification of these products was facilitated by use of reversed phase cartridges.