| 摘要 |
<p>A method for detecting PCR amplification of a target DNA molecule in a sample is described which employs a forward PCR primer having a sequence that is complementary to the target double stranded nucleic acid and a tail sequence, a reverse PCR primer having a sequence that is complementary to the target double stranded nucleic acid, and a dual labelled probe containing an oligonucleotide sequence identical to the tail sequence of the forward PCR primer oligonucleotide and a reporter label and a quencher-label separated by a nuclease susceptible cleavage site. The method comprises the steps of incubating the forward and reverse PCR primers and dual labelled probe together, performing at least two rounds of PCR to generate a double stranded nucleic acid comprising the tail region and a sequence complementary to the tail region, and initiating a further round of PCR in which the oligonucleotide probe binds to the sequence complementary to the tail region, whereby the oligonucleotide probe is displaced and cleaved (e.g. by exonuclease activity of the DNA polymerase) resulting in a detectable signal from the reporter label. Associated methods for detecting different alleles and kits comprising the aforementioned primers and probes are also claimed.</p> |
