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US 2014/0178882 A1Published application
Nucleic acid binding dyes and uses therefor
AssigneePromega Corporation
InventorsMark McDougall, Stephen Dwight
Life of this patent
Estimating the term…
Abstract
The invention provides novel compounds and compositions of Formulas I and II, as well as methods of using them. The compounds can be used, for example, to quantify an amount of double stranded DNA in a sample subjected to nucleic acid amplification, or for real time monitoring of a nucleic acid amplification reaction. The compounds can be provided in a kit, for example, with other reagents and instructions for using the compounds and reagents.
Claims16
A method to quantify the amount of a nucleic acid in a sample subjected to nucleic acid amplification, comprising:
- a) contacting the sample with a compound according to Formula I:
- wherein
- R1, R2 and R3 are each independently H, alkyl, cycloalkyl, aryl, heteroaryl, amino, hydroxy, or halo;
- or R1 and R2, or R2 and R3, taken together with the atoms to which they are attached form a fused benzo ring that is optionally substituted with 1, 2, 3, or 4 alkyl, cycloalkyl, aryl, heteroaryl, amino, hydroxy, or halo groups;
- Y is S, O, CF2, or NR4;
- R4 is (C1-C8)alkyl, aryl, or (C1-C8)alkaryl;
- W taken together with the atoms to which it is attached is a 5-, 6-, 7-, or 8-membered heterocyclic ring;
- n is 0, 1, or 2;
- Q is Q1 or Q2:
- wherein
- Y is —CR13═CR14—;
- m and p are 0 or 1 such that m+p=1;
- R6 is (C1-C8)alkyl, aryl, or (C1-C8)alkaryl;
- R7, R8, R9, R10, R11, and R12 are each independently H, alkyl, cycloalkyl, aryl, heteroaryl, amino, or halo;
- one of R13 and R14 is H and the other is NR15R16;
- R15 is (C1-C6)alkyl or (C1-C6)alkyl-NR17R18, wherein R17 and R18 are each independently H or (C1-C6)alkyl;
- and
- R16 is (C1-C6)alkyl-NR12R18, wherein R12 and R18 are each independently H or (C1-C6)alkyl;
- or a salt thereof;
- b) detecting fluorescence in the amplification reaction;
- and
- c) correlating the amount of fluorescence in the amplification reaction with the amount of nucleic acid in the reaction.
19 more elements
A method to quantify the amount of a nucleic acid in a sample subjected to nucleic acid amplification, comprising:
- a) amplifying a nucleic acid in a sample, the sample comprising a target nucleic acid molecule, one or more nucleic acid primers specific for target sequences in the nucleic acid to be amplified, a polymerase, and a compound according to Formula I, so as to yield a mixture comprising amplified nucleic acid:
- wherein
- R1, R2 and R3 are each independently H, alkyl, cycloalkyl, aryl, heteroaryl, amino, hydroxy, or halo;
- or R1 and R2, or R2 and R3, taken together with the atoms to which they are attached form a fused benzo ring that is optionally substituted with 1, 2, 3, or 4 alkyl, cycloalkyl, aryl, heteroaryl, amino, hydroxy, or halo groups;
- Y is S, O, CF2, or NR4;
- R4 is (C1-C8)alkyl, aryl, or (C1-C8)alkaryl;
- W taken together with the atoms to which it is attached is a 5-, 6-, 7-, or 8-membered heterocyclic ring;
- n is 0, 1, or 2;
- Q is Q1 or Q2:
- wherein
- Y is —CR13═CR14—;
- m and p are 0 or 1 such that m+p=1;
- R6 is (C1-C8)alkyl, aryl, or (C1-C8)alkaryl;
- R7, R8, R9, R10, R11, and R12 are each independently H, alkyl, cycloalkyl, aryl, heteroaryl, amino, or halo;
- one of R13 and R14 is H and the other is NR15R16;
- R15 is (C1-C6)alkyl or (C1-C6)alkyl-NR17R18, wherein R17 and R18 are each independently H or (C1-C6)alkyl;
- and
- R16 is (C1-C6)alkyl-NR17R18, wherein R17 and R18 are each independently H or (C1-C6)alkyl;
- or a salt thereof;
- b) detecting fluorescence in the mixture;
- and
- c) correlating the amount of fluorescence in the mixture with the amount of nucleic acid in the mixture.
19 more elements
A method of real time monitoring of a nucleic acid amplification reaction, comprising
- a) amplifying a target nucleic acid sequence in a target nucleic acid molecule in reaction mixture containing a sample having the target nucleic acid molecule, one or more nucleic acid primers specific for sequences in the target, and a compound according to Formula I:
- wherein
- R1, R2 and R3 are each independently H, alkyl, cycloalkyl, aryl, heteroaryl, amino, hydroxy, or halo;
- or R1 and R2, or R2 and R3, taken together with the atoms to which they are attached form a fused benzo ring that is optionally substituted with 1, 2, 3, or 4 alkyl, cycloalkyl, aryl, heteroaryl, amino, hydroxy, or halo groups;
- Y is S, O, CF2, or NR4;
- R4 is (C1-C8)alkyl, aryl, or (C1-C8)alkaryl;
- W taken together with the atoms to which it is attached is a 5-, 6-, 7-, or 8-membered heterocyclic ring;
- n is 0, 1, or 2;
- Q is Q1 or Q2:
- wherein
- Y is —CR13═CR14—;
- m and p are 0 or 1 such that m+p=1;
- R6 is (C1-C8)alkyl, aryl, or (C1-C8)alkaryl;
- R7, R8, R9, R10, R11, and R12 are each independently H, alkyl, cycloalkyl, aryl, heteroaryl, amino, or halo;
- one of R13 and R14 is H and the other is NR15R16;
- R15 is (C1-C6)alkyl or (C1-C6)alkyl-NR17R18, wherein R17 and R18 are each independently H or (C1-C6)alkyl;
- and
- R16 is (C1-C6)alkyl-NR17R18, wherein R17 and R18 are each independently H or (C1-C6)alkyl;
- or a salt thereof;
- b) illuminating the reaction mixture with light of a selected wavelength that is absorbed by the compound;
- and
- c) detecting the fluorescence emission in the reaction mixture.
19 more elements
Dependencies are inferred from the claim text.