The production of PCR products with 5′ single-stranded tails using primers that incorporate novel phosphoramidite intermediates

Abstract

We have prepared several novel phosphoramidltes and have synthesised ollgonucleotldes incorporating them internally. The presence of these residues in an ollgonucleotlde template presents an impassible barrier to primed synthesis by Taq DNA polymerase. When extended as polymerase chain reaction products, these ollgonucleotldes no longer serve as templates for the polymerase beyond the insertion sites of the modified intermediates, thereby producing single-stranded tails on amplification products. These tails can then be used for solid phase capture and colourlmetric detection of PCR products.