Patent ID: 7361489
Filing Date: 2008-04-22
Classification: C12Q

Abstract:
1. A method for detecting a target nucleic acid in a nucleic acid containing sample comprising: (a) contacting the nucleic acid containing sample with a circular oligonucleotide probe under conditions that allow hybridization between complementary sequences in the target nucleic acid and the circular oligonucleotide probe; (b) adding at least one forward primer comprising a sequence complementary to a portion of the circular oligonucleotide probe, under conditions where the forward primer is extended around the circular oligonucleotide probe for multiple revolutions to form a single-stranded DNA molecule comprising repeating units complementary to the sequence of the circular probe; (c) adding at least one oligonucleotide primer pair comprising a first primer and a second primer, wherein (d) adding at least one reverse primer comprising a sequence that is substantially identical to a portion of the circular oligonucleotide probe and wherein the forward primer, reverse primer, and the first primer and the second primer of the oligonucleotide primer pair are not identical; (e) adding a DNA polymerase having strand displacement activity and lacking 3′ to 5′ exonuclease activity; and (f) amplifying the circular oligonucleotide probe using ramification-extension amplification method (RAM) thus producing an amplification product comprising a sequence that is substantially identical to a sequence in the circular probe, and separating the signal generating moiety selected from the group consisting of a fluorescent agent and a chemiluminescent agent from the quenching, masking or inhibitory moiety by the action of the DNA polymerase having strand displacement activity and lacking 3′ to 5′ exonuclease activity during the amplification method, wherein detection of the signal indicates the presence of the target nucleic acid in the nucleic acid containing sample.