Patent ID: 6255051
Filing Date: 2001-07-03
Classification: C12Q

Abstract:
A method of detecting a target polynucleotide sequence in a biological sample comprising multiple sequential polynucleotide displacement for signal generation comprising the following steps:(a) forming a first probe complex by contacting a first polynucleotide sequence comprising a complementary target binding region (CTBR), with a second polynucleotide sequence comprising a partial target binding region (PTBR) and a cascade binding region (CBR), under conditions suitable for hybridization between said first and said second polynucleotide sequences, thereby forming a first probe complex;(b) forming a first displacement complex by contacting the complex of (a) with a target polynucleotide sequence comprising a target binding region (TBR), under conditions suitable for said target polynucleotide sequence to displace said second polynucleotide sequence and hybridize to said first polynucleotide sequence, thereby forming a first displacement complex and a displaced second polynucleotide sequence;(c) forming a second probe complex by contacting a fourth polynucleotide sequence comprising a complementary cascade binding region (CCBR), with a fifth polynucleotide sequence comprising at least one cascade binding region (CBR), under conditions suitable for hybridization between said fourth and fifth polynucleotide sequences, thereby forming a second probe complex; and(d) forming a second displacement complex by contacting the displaced second polynucleotide sequence of (b) with the second probe complex of (c) under conditions suitable for the displaced second polynucleotide sequence of (b) to displace said fifth polynucleotide sequence of (c) and hybridize with said fourth polynucleotide sequence of (c),thereby producing a second displacement complex and a liberated fifth polynucleotide sequence which generates at least one signal, and wherein detection of the signal is indicative of the presence of the target polynucleotide sequence in a biological sample.