| US 7,413,857 B2 | ||
| Methods for using riboprimers for strand displacement replication of target sequences | ||
| Gary A. Dahl, Madison, Wis. (US); Jerome J. Jendrisak, Madison, Wis. (US); and Agnes J. Radek, Madison, Wis. (US) | ||
| Assigned to Epicentre Technologies, Madison, Wis. (US) | ||
| Filed on Nov. 21, 2003, as Appl. No. 10/719,168. | ||
| Claims priority of provisional application 60/428013, filed on Nov. 21, 2002. | ||
| Prior Publication US 2004/0180361 A1, Sep. 16, 2004 | ||
| Int. Cl. C12Q 1/68 (2006.01); C12P 19/34 (2006.01); C07H 21/04 (2006.01) | ||
| U.S. Cl. 435—6 [435/91.2; 536/24.33] | 64 Claims |
| 1. A method for amplifying a target nucleic acid sequence comprising a target nucleic acid, the method comprising the steps
of:
a) hybridizing a riboprimer to a DNA template comprising the target nucleic acid sequence, wherein said riboprimer comprises:
i) only ribonucleotides, or ii) only purine ribonucleotides and only pyrimidine nucleotides, wherein at least one of the pyrimidine
nucleotides is a pyrimidine 2′-deoxyribonucleotide having a non-canonical substituent, which substituent is neither an H nor
an OH, on the 2′-position of the deoxyribose sugar moiety;
b) extending the riboprimer with a DNA polymerase that lacks 5′-to-3′ exonuclease activity; and
c) cleaving the annealed riboprimer with an RNAse H enzyme such that another riboprimer hybridizes to the template and repeats
primer extension, whereby multiple copies of the complementary sequence of the target nucleic acid sequence are produced.
|