PROTECTING FULLY FUNCTIONAL NUCLEOTIDES FROM DEGRADATION
This application relates to methods and compositions for protecting fully functional nucleotides (ffNs) against degradation. In some examples, micelles or surfactants are used to protect ffNs. In some examples, bulky cation compounds are used to protect ffNs. In some examples, the compositions that protect ffNs are included in lyophilized material.
1 . A method of stabilizing fully functional nucleotides (ffNs), the method comprising:
including the ffNs and at least one surfactant into lyophilized material.
2 . The method of claim 1 , wherein a concentration of the ffNs is between about 10 μM and about 15,000 μM.
3 . The method of claim 1 , wherein a concentration of the at least one surfactant is between about 0.5% and about 2.5%.
4 . The method of claim 1 , wherein including the ffNs and the at least one surfactant into the lyophilized material comprises freeze-drying the ffNs.
5 . The method of claim 1 , wherein the ffNs are encapsulated in a hydrophobic core of the surfactant, which inhibits or prevents hydrolysis of the ffNs.
6 . The method of claim 1 , wherein the at least one surfactant comprises any one or more of tricosaethylene glycol dodecyl ether, poloxamer 407, polyethylene glycol (PEG) stearate, SB-12, lauramidopropyl betaine, n-dodecyl-β-D-maltoside, CHAPS, polyoxyethylene sorbitan monolaurate, and polyoxyethylene sorbitan monopalmitate.
7 . A method of stabilizing a fully functional nucleotides (ffNs), the method comprising:
including the ffNs and at least one bulky cation compound into lyophilized material.
8 - 17 . (canceled)
18 . A method of preparing fully functional nucleotides (ffNs) for sequencing, the method comprising:
mixing the ffNs with at least one surfactant in a solution;
diluting a concentration of the ffNs in a sequencing mix relative to a concentration of the ffNs in the solution by a factor of between about 10× and about 200×; and
diluting a concentration of the at least one surfactant in the sequencing mix, relative to a concentration of the at least one surfactant in the solution, by a factor of between about 10× and about 1,000×.
19 . The method of claim 18 , wherein the concentration of the ffNs in the solution is between about 10 μM and about 15,000 μM, and the concentration of the ffNs in the sequencing mix is between about 5 μM and about 9 μM.
20 . The method of claim 18 , wherein the concentration of the at least one surfactant in the solution is between about 1.5% and about 2.5%, and the concentration of the at least one surfactant in the sequencing mix is between about 0.05% and about 0.25%.
21 . The method of claim 18 , wherein the at least one surfactant comprises any one or more of tricosaethylene glycol dodecyl ether, poloxamer 407, CHAPS, polyoxyethylene sorbitan monolaurate, and polyoxyethylene sorbitan monopalmitate.
22 . The method of claim 18 , further comprising performing a sequencing reaction through incorporating the ffNs onto a nucleic acid.
23 . The method of claim 22 , wherein the sequencing reaction comprises a sequencing by synthesis reaction.
24 . A method of preparing ffNs for sequencing, the method comprising:
mixing the ffNs with at least one bulky cation compound in a solution;
diluting a concentration of the ffNs in a sequencing mix relative to a concentration of the ffNs in the solution by a factor of between about 10× and about 200×; and
diluting the concentration the at least one bulky cation compound in the sequencing mix relative to the concentration of the at least one bulky cation compound in the solution, by a factor of between about 100× and about 1,000×.
25 - 29 . (canceled)