Homogeneous polynucleotide displacement assay method kit and reagent complex

DETAILED DESCRIPTION OF THE INVENTION The basic elements of the reagent complex provided by the present invention and used in the method and kit of the present invention are a probe polynucleotide and a signal strand, which as described below, may either be bound to each other only by complimentary base pairing or which may be further covalently attached by phosphate/sugar polynucleotide backbone (or otherwise be further attached covalently or non-covalently).
The probe polynucleotide has a target binding region complementary to the target nucleotide sequence which will be determined.
As described more fully in U.
S.
application Ser.
No.
607,885, the target binding region may be perfectly complementary to the target nucleotide sequence or may contain a limited number of mismatches.
Furthermore, the target binding region is conveniently divided into a portion called the Signal Strand Binding Region (or Labeled Polynucleotide Bindnng Region and thus LBR in the figures) to which a portion of the signal strand is bound by complimentary base pairing in the reagent complex.
As illustrated by FIG.
1G of U.
S.
Ser.
No.
607,885, a small number of additional bases of the signal strand may be bound to a small portion of the probe polynucleotide outside the target binding region (called the Residual Binding Region or RBR) but such residual binding region is preferably not present.
A further portion (or portions) usually present in the target binding region of the probe polynucleotide is in single-stranded form in the reagent complex and is referred to as the initial binding region (IBR) in that the target nucleotide sequence can bind initially in this region prior to any displacement of nucleotides of the signal strand.
As described in U.
S.
Ser.
No.
607,885, the size of the target binding region is not independently critical, but can be thought as the sum of preferred or more preferred lengths of LB and IBR.
The signal strand binding region (LBR) is preferably at least 25 nucleotides in length, more preferably 50-1000 nucleotides in length and most preferably 300-1000 nucleotides in length