Patent ID: 9334533
Filing Date: 2016-05-10
CPC Classification: C12Q

Claim Text:
1. A method for detecting the presence of a variation or the type of a variation in a target nucleic acid, comprising: (1) amplifying a fragment comprising the nucleic acid to be tested using asymmetric PCR after adding a probe to the amplification reaction mixture for the asymmetric PCR, wherein one PCR amplification primer in the reaction mixture is relatively in excess and the strand produced with the elongation of said primer hybridizes with the probe and the polymerase used in the asymmetric PCR has an exonuclease activity; wherein the probe is a self-quenched nucleic acid probe, wherein the probe is labeled at opposite ends with a fluorescent group and a quenching group in such a way that fluorescence or fluorescence intensity increases when the probe hybridizes with the target nucleic acid sequence compared to fluorescence or fluorescence intensity in the absence of the target nucleic acid sequence, wherein: if the 5′ end of the probe is labeled with the fluorescent group, then the 3′ end of the probe is labeled with the quenching group; or if the 3′ end of the probe is labeled with the fluorescent group, then the 5′ end is labeled with the quenching group, and said probe does not comprise a modification that is able to resist the exonuclease activity of a polymerase; and wherein the probe is a hairpin structure probe; and (2) determining whether the target nucleic acid has a sequence variation compared to a reference or wild-type nucleic acid by melting curve analysis of the amplification product.