| 摘要 |
An oligonucleotide probe is disclosed, the probe including an oligonucleotid e, a fluorescer molecule attached to a first end of the oligonucleotide and a quencher molecule attached to the opposite end of the oligonucleotide. The probe is rendered impervious to digestion by the 5'3' exonuclease activity o f a polymerase and the 5'3' extension by a polymerase. The invention also includes methods for performing combined PCR amplification and hybridization probing, one such method including the steps of contacting a target nucleic acid sequence with PCR reagents and an oligonucleotide probe as described above, and subjecting these reagents to thermal cycling. One preferred refinement of the above method further includes the addition of a strand displacer to facilitate amplification. Additional similar combined PCR hybridization methods are disclosed, such methods not requiring probes havin g their 5' ends protected, wherein (i) the polymerase lacks 5'3' exonuclease activity, (ii) a 5'3' exonuclease inhibitor is included, and (iii) an exonuclease deactivation step is performed.
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