Systems and methods for nucleic acid sequencing
The present disclosure provides methods and systems for sequencing nucleic acid molecules. Methods may include sequencing double-stranded nucleic acids or single-stranded nucleic acids. Sequencing may include the use of nucleotides coupled to electrostatic moieties. The electrostatic moieties may be detected by a sensor array. The electrostatic moieties may be reversible electrostatic moieties that are cleaved from the nucleic acid molecule after incorporation of the nucleotide. The electrostatic moieties may be irreversible electrostatic moieties. Nucleotides comprising irreversible electrostatic moieties may be incorporated into the nucleic acid molecule, detected by the sensor array, and exchanged for non-detectable nucleotides.
1. A method for processing or analyzing a nucleic acid molecule, comprising:
(a) providing a sensor that is electrically coupled to a charge double layer comprising at least a portion of said nucleic acid molecule;
(b) bringing said nucleic acid molecule in contact with a nucleotide coupled to an effector in presence of a polymerizing enzyme under conditions sufficient to incorporate said nucleotide into a growing strand complementary to said nucleic acid molecule, wherein said effector is capable of modulating charge distribution at said nucleotide;
(c) subsequent to (b), detecting one or more signals associated with said charge double layer with said sensor, which one or more signals are indicative of said nucleotide coupled to said effector having been incorporated into said growing strand, wherein said effector modulates charge distribution at said nucleotide during said detecting; and
(d) reducing effects of said effector on said charge distribution at said nucleotide.
2. The method of claim 1 , wherein said nucleic acid molecule is coupled to a bead.
3. The method of claim 1 , wherein said nucleic acid molecule is coupled to a surface of said sensor.
4. The method of claim 1 , wherein said nucleic acid molecule is among a plurality of nucleic acid molecules having sequence homology with one another, and wherein said charge double layer comprises said plurality of nucleic acid molecules.
5. The method of claim 1 , wherein said sensor is part of a sensor array.
6. The method of claim 1 , wherein said effector is coupled to a nucleobase of said nucleotide.
7. The method of claim 1 , wherein said effector is an electrostatic moiety.
8. The method of claim 1 , wherein (d) comprises removing said effector from said nucleotide.
9. The method of claim 1 , wherein said nucleotide comprises a reversible terminator that prevents an additional nucleotide from stably hybridizing to said nucleic acid molecule.
10. The method of claim 9 , further comprising removing said reversible terminator subsequent to (c) and prior to incorporation of another nucleotide into said growing strand.
11. The method of claim 1 , wherein said nucleotide is among a plurality of nucleotides of different types, and wherein (b) comprises bringing said nucleic acid molecule in contact with said plurality of nucleotides.
12. The method of claim 11 , wherein said plurality of nucleotides are reversibly coupled to a single type of effector.
13. The method of claim 11 , wherein said plurality of nucleotides are reversibly coupled to different types of effectors.
14. The method of claim 1 , wherein said one or more signals are steady state signals.
15. The method of claim 1 , wherein said one or more signals are electrical signals generated by an impedance or change in impedance in said charge double layer.
16. The method of claim 1 , wherein said one or more signals are electrical signals generated by a conductivity or change in conductivity in said charge double layer.
17. The method of claim 1 , wherein said one or more signals are electrical signals generated by charge or change in charge in said charge double layer.
18. The method of claim 1 , wherein said sensor comprises a plurality of electrodes in contact with said charge double layer.
19. The method of claim 1 , wherein (b)-(d) are repeated until a sequence or a length of said nucleic acid molecule is determined.