Patent ID: 7387887

Claim:
A PCR reaction mixture comprising a. a target nucleic acid, b. PCR reagents, c. a pair of oligonucleotide primers configured for amplifying a portion of the target nucleic acid to produce an amplicon, and d. a dsDNA binding dye having the formula: wherein the moiety Y represents an optionally-substituted fused monocyclic or polycyclic aromatic ring or an optionally-substituted fused monocyclic or polycyclic nitrogen-containing heteroaromatic ring; X is oxygen, sulfur, selenium, tellurium or a moiety selected from C(CH 3 ) 2 and NR 1 , where R 1 is hydrogen or C 1-6 alkyl; R 2 is selected from the group consisting of C 1-6 alkyl, C 3-8 cycloalkyl, aryl, aryl(C 1-3 alkyl), hydroxyalkyl, alkoxyalkyl, aminoalkyl, mono and dialkylaminoalkyl, trialkylammoniumalkyl, alkylenecarboxylate, alkylenecarboxamide, alkylenesulfonate, optionally substituted cyclic heteroatom-containing moieties, and optionally substituted acyclic heteroatom-containing moieties; t=0 or 1; Z is a charge selected from 0 or 1; R 3 , R 9 , and R 10 are each independently selected from the group consisting of hydrogen, C 1-6 alkyl, and arylcarbonyl; n=0,1, or 2; and Q is an heterocycle selected from the group of structures consisting of: wherein R 5 , R 6 , R 7 , and R 8 are independently selected from the group consisting of hydrogen, halogen, alkyl, cycloalkyl, heteroalkyl, heterocycloalkyl, alkenyl, polyalkenyl, alkynyl, polyalkynyl, alkenylalkynyl, aryl, heteroaryl, alkoxy, alkylthio, arylthio, arylcarbonylthio, dialkylamino, cycloheteroalkylcarbonylthio, dialkylaminoalkylcarbonylthio, cycloalkylthio, cycloheteroalkylthio, trialkylanimoniumalkylthio, and nucleosidylthio, each of which may be optionally substituted; an acyclic heteroatom-containing moiety or a cyclic heteroatom-containing moiety, a BRIDGE-DYE, and a reactive group, each of which optionally includes a guaternary ammonium moiety, and R 4 is selected from the group consisting of arylcarbonylthio, cycloheteroalkylcarbonylthio, dialkylaminoalkylcarbonylthio, cycloalkylthio, cycloheteroalkylthio, trialkylammoniumalkylthio, and nucleosidylthio, each of which may be optionally substituted, wherein the dye is selected from the group consisting of F7, N7, O7, P7, Q7, R7, S7, T7, V7, W7, X7, T8, A9, C9, I9, J9, K9, L9, M9, N9, O9, P9 and R9 as presented in Table 2.