Methods of using dyes in association with nucleic acid staining or detection
First Claim
1. A method of detecting the presence or absence of a nucleic acid in a sample, comprising:
- combining a sample comprising a nucleic acid with at least one fluorescent nucleic acid binding dye having the formula;
1 Assignment
0 Petitions
Accused Products
Abstract
Methods of using dyes and associated technology are provided. A dye, such as a monomeric dye or a dimeric dye, may be used in a nucleic acid gel staining application and/or a nucleic acid detection application. A dimeric dye, such as a dimeric dye capable of forming a hairpin-like structure, may be used to stain and/or detect nucleic acids via a release-on-demand mechanism. A dimeric dye having low background fluorescence in the absence of nucleic acids and high fluorescence in the presence of nucleic acids, upon binding therewith, may be used to stain and/or detect nucleic acids. Buffers comprising a weak acid and a salt of the weak acid, such as a lithium salt, are also provided to allow for effective prestaining of nucleic acids.
47 Citations
29 Claims
-
1. A method of detecting the presence or absence of a nucleic acid in a sample, comprising:
combining a sample comprising a nucleic acid with at least one fluorescent nucleic acid binding dye having the formula;
-
2. The method of claim 1, wherein said running buffer comprises at least one weak acid and a lithium salt of said at least one weak acid.
-
3. The method of claim 1, wherein said sample further comprises a loading dye.
-
4. The method of claim 3, wherein said loading dye is xylene cyanol, cresol red, bromophenol blue, orange G or tartrazine.
-
5. The method of claim 1, wherein Q1 and Q2 are different.
-
6. The method of claim 1, wherein when Q1 and/or Q2 is a phenanthridinium dye it has the structure of Formula I or Formula II:
-
7. The method of claim 6, wherein said phenanthridinium dye has the structure of Formula I, R1 is a phenyl and R2 is -BRIDGE-Q1 or -BRIDGE-Q2.
-
8. The method of claim 7, wherein both Q1 and Q2 are the same phenanthridinium dye.
-
9. The method of claim 6, wherein said phenanthridinium dye has the structure of Formula I, R2 is a phenyl and R1 is -BRIDGE-Q1 or -BRIDGE-Q2.
-
10. The method of claim 9, wherein both Q1 and Q2 are the same phenanthridinium dye.
-
11. The method of claim 1, wherein when Q1 and/or Q2 is a acridinium dye it has the structure of Formula III:
-
12. The method of claim 11, wherein R1 and R2 are H;
- R3 is -BRIDGE-Q1 or -BRIDGE-Q2; and
R4 and R5 are —
CH3.
- R3 is -BRIDGE-Q1 or -BRIDGE-Q2; and
-
13. The method of claim 12, wherein both Q1 and Q2 are the same acridinium dye.
-
14. The method of claim 1, wherein when Q1 and/or Q2 is a asymmetric cyanine dye it has the structure of Formula IV:
-
15. The method of claim 14, wherein R1 is —
- CH3;
R2 is H;
n is 0;
R3 is H;
R4 is -BRIDGE-Q1 or -BRIDGE-Q2; and
R5 and R6 together with the carbon atoms they are attached to form a fused benzene ring.
- CH3;
-
16. The method of claim 15, wherein both Q1 and Q2 are the same asymmetric cyanine dye.
-
17. The method of claim 1, wherein said weak acid has a pKa of about 8.0 to about 10.5.
-
18. The method of claim 1, wherein said weak acid is boric acid.
-
19. The method of claim 1, wherein the gel buffer or running buffer has a pH of about 7.7 to about 9.0.
-
20. The method of claim 19, wherein the gel buffer or running buffer has a pH of about 8 to about 8.8.
-
21. The method of claim 1, wherein said gel buffer or running buffer comprises from about 0.5 mM to about 30 mM Li .
-
22. The method of claim 1, wherein said gel buffer or running buffer further comprises one or more of a magnesium chelator, a detergent, or a preservative.
-
23. The method of claim 1, wherein said fluorescent nucleic acid binding dye has the structure:
-
24. The method of claim 1, wherein said fluorescent nucleic acid binding dye has the structure:
-
25. The method of claim 1, wherein said fluorescent nucleic acid binding dye has the structure:
-
26. The method of claim 1, wherein said fluorescent nucleic acid binding dye has the structure:
-
27. The method of claim 1, wherein said fluorescent nucleic acid binding dye has the structure:
-
28. The method of claim 1, wherein said fluorescent nucleic acid binding dye has the structure:
-
29. The method of claim 1, wherein said fluorescent nucleic acid binding dye has the structure:
Specification