Efficient methods for attachment of thiol specific probes to the 3′-ends of synthetic oligodeoxyribonucleotides

Methodology is described for the synthesis of DNA oligomers containing a free 3′-thiol group which can be selectively crosslinked with a wide variety of probes. This chemistry is compatible with both phosphotriester and phosphoramidite solid phase chemistry. Moreover, the sulphydryl group is introduced into the 3′-nucleoside solid support linkage prior to oligonucleotide synthesis. Consequently, no additional coupling steps are required after oligonucleotide synthesis, and isolation of the 3′-thiol oligonucleotide requires only one additional deprotection step. Cross-linking of the thiol-containing oligonucleotide to a fluorescent probe was carried out with high selectivity, in high yield, and under mild conditions.