Patent ID: 8911948

Claim:
A method of amplifying a target DNA sequence, said method comprising the steps of: (a) providing a reaction mixture comprising (i) an oligonucleotide primer having a cleavage domain with a cleavage site, located within or adjacent to the cleavage domain, which is cleavable by an RNase H2 enzyme, wherein said cleavage site is positioned 5′ of a blocking group at or near the 3′-end of the oligonucleotide primer which prevents primer extension and/or PCR, (ii) a sample nucleic acid that may or may not have the target sequence, (iii) a DNA polymerase, (iv) an RNase H2 enzyme encoded by SEQ ID NO:4, and (v) optionally a second primer in reverse orientation to support PCR; (b) hybridizing the blocked primer to the target DNA sequence to form a double-stranded substrate; (c) cleaving the hybridized blocked primer with said RNase H2 enzyme to remove the blocking group from the primer; and (d) extending the cleaved primer with the DNA polymerase.