Methods of attaching unprotected oligonucleotides to DNA-binding, fluorescent, or reactive ligands for synthesis of antisense or gene-directed agentsand probes
As. Boutorine et al., Methods of attaching unprotected oligonucleotides to DNA-binding, fluorescent, or reactive ligands for synthesis of antisense or gene-directed agentsand probes, MOL BIOL, 34(6), 2000, pp. 804-813
The article describes the optimized methods for covalent attachment of unpr
otected oligonucleotides to functionally important ligands through their te
rminal phosphate or thiophosphate, including covalent attachment of one oli
gonucleotide to another. A comparative description of methods is presented
for selective introduction of the phosphate, thiophosphate, amino, sulfhydr
yl, aldehyde, carboxylic, and other groups into the terminal nucleotide usi
ng chemical and enzymatic reactions both in aqueous and organic media. Depe
nding on their chemical nature, these groups can then interact with electro
philic or nucleophilic ligands carrying aliphatic or aromatic amino groups,
hydrazido, sulfhydryl, disulfide, carboxylic, hydroxyl, aldehyde, bromo- o
r iodoalkyl, isothiocyanate, and other functions. The available methods all
ow one to vary the size of the linker between the oligonucleotide and ligan
d, its hydrophobicity and stability in acidic or alkaline media. The use of
the disulfide bond permits cleavage of the oligonucleotide-ligand linkage
in mild conditions.