Method for nucleotide detection
A method of inhibiting light-induced degradation of nucleic acids includes irradiating a portion of the nucleic acids in the presence of a detection solution comprising a polyphenolic compound. A method of detecting a nucleic acid having a fluorescent tag includes irradiating at least a portion of the nucleic acid with light of a suitable wavelength to induce a fluorescence emission and detecting the fluorescence emission. Optionally, the polyphenolic compound is gallic acid, a lower alkyl ester thereof, or mixtures thereof. A kit includes one or more nucleotides, an enzyme capable of catalyzing incorporation of the nucleotides into a nucleic acid strand and a polyphenolic compound suitable for preparing a detection solution.
1. A method of inhibiting light-induced degradation of nucleic acids during a detection step, the method comprising:
introducing an incorporation solution into a flow cell, said incorporation solution comprising polymerase and a fluorescently tagged nucleotide, said flow cell comprising an array of nucleic acids attached to a support;
replacing the incorporation solution with a detection solution comprising gallic acid, a lower alkyl ester thereof, or mixtures thereof, and further comprising urea,
irradiating a portion of said nucleic acids in the presence of said detection solution, wherein said detection solution reduces the amount of light-induced degradation of said nucleic acids.
2. The method of claim 1 , wherein said gallic acid, said lower alkyl ester thereof, or said mixtures thereof is present in a concentration ranging from between about 10 mM to about 200 mM.
3. The method of claim 1 , further comprising adding an additional fluorescently tagged nucleotide to said array and repeating said detection step in a cycle.
4. The method of claim 3 , comprising repeating said adding and detection steps for at least 50, 75, or 100 cycles.
5. The method of claim 3 , comprising repeating said adding and detection steps for a number of cycles in a range from between about 100 cycles to about 1,000 cycles.
6. The method of claim 1 , wherein the presence of said detection solution reduces a detection error rate by greater than 20% relative to a control.
7. The method of claim 1 , wherein said irradiation step is conducted in a range from about 360 nm to about 700 nm.
8. The method of claim 1 , wherein said irradiation step is conducted with a light source having power in a range between about 5 to about 500 milliwatts.
9. The method of claim 1 , wherein said irradiation step is conducted for a time period of about 0.1 seconds to about 10 minutes.
10. The method of claim 1 , wherein said array comprises a primer template.
11. A method of detecting a nucleic acid having a fluorescent tag comprising:
a) introducing an incorporation solution into a flow cell, said incorporation solution comprising polymerase and a fluorescently tagged nucleotide, said flow cell comprising an array of nucleic acids attached to a support, to add a fluorescently tagged nucleotide to said nucleic acid;
b) replacing the incorporation solution with a detection solution;
c) irradiating at least a portion of said nucleic acid with light in the presence of the detection solution, wherein said light comprises a suitable wavelength to induce a fluorescence emission;
d) detecting said fluorescence emission; and
e) repeating steps a) through d);
wherein the detection solution comprises gallic acid, a lower alkyl ester thereof, or mixtures thereof, and further comprising urea, said detection solution inhibiting light-induced degradation of said nucleic acid.
12. The method of claim 11 , wherein said gallic acid, said lower alkyl ester thereof, or said mixtures thereof is present in a concentration ranging from between about 10 mM to about 200 mM.
13. The method of claim 11 , comprising at least 50, 75, or 100 cycles repeating step e.
14. The method of claim 11 , wherein the presence of said detection solution reduces a detection error rate by greater than 20% relative to a control.
15. The method of claim 11 , wherein said irradiation step is conducted in a range from about 360 nm to about 700 nm.
16. The method of claim 11 , wherein said irradiation step is conducted with a light source having power in a range between about 5 to about 500 milliwatts.
17. The method of claim 11 , wherein said irradiation step is conducted for a time period of about 0.1 seconds to about 10 minutes.
18. The method of claim 11 , wherein step a) comprises using a polymerase to add a single nucleotide.
19. The method of claim 11 , wherein said array comprises a primer template.