Polymerization of nucleic acids using activation by polyphosphorolysis (APP) reactions

摘要

This disclosure relates to methods of performing activation by polyphosphorolysis (APP) reactions using at least one of the polyphosphorylating agents triphosphate, polyphosphate, imidodiphosphate, thiodiphosphate (or μ-monothiopyrophosphate), and related compounds.

主权项

1. A method for producing an unblocked oligonucleotide from a blocked oligonucleotide comprising a non-extendable 3′ end removable by polyphosphorolysis, the method comprising
contacting the blocked oligonucleotide hybridized to a target nucleic acid with one or more polyphosphorolyzing agents selected from the group consisting of:
a compound of the general formula I: wherein n is selected from the group consisting of 1, 2, 3, 4, 5, 6, 7, 8, 9, and 10; anda compound of general formula II: wherein n and/or m are selected from the group consisting of 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, and 10, with the proviso that n and m cannot both be 0, and
X is selected from the group consisting of: in the presence of an enzyme having polyphosphorolysis activity and removing the 3′ terminal nucleotide of the blocked oligonucleotide to produce the unblocked oligonucleotide.