Methods and apparatus for characterizing polynucleotides
Systems and methods for analysis of polymers, e.g., polynucleotides, are provided. The systems are capable of analyzing a polymer at a specified rate. One such analysis system includes a structure having a nanopore aperture and a molecular motor, e.g., a polymerase, adjacent the nanopore aperture.
1. An analysis system comprising:
a structure having a nanopore aperture that separates two independent adjacent pools of solution;
a helicase that is capable of moving a polynucleotide with respect to the nanopore aperture at a rate of less than 250 Hz and that is located in at least one of the pools; and
a detector capable of measuring transport properties of the polynucleotide as it moves with respect to the nanopore.
2. The system of claim 1 , wherein the nanopore aperture is solid-state.
3. The system of claim 1 , wherein the nanopore aperture comprises a biological nanopore.
4. The system of claim 1 , wherein the transport properties are the amplitude or duration of individual conductance or electron tunneling current changes across the nanopore aperture.
5. The system of claim 1 , wherein the helicase is selected from the group consisting of E - coli bacteriophage T7 gp4 and T4 gp41 gene proteins, and the E. coli proteins DnaB, RuvB, and rho.
6. The system of claim 1 , wherein the detector comprises electrodes at or near the nanopore aperture.
7. The system of claim 1 , wherein the detector comprises electrodes placed within the adjacent pools.
8. The system of claim 1 , wherein the rate of the helicase is less than or equal to 200 Hz.
9. The system of claim 1 , wherein the rate of the helicase is less than or equal to 150 Hz.
10. The system of claim 1 , wherein the helicase is capable of moving the polynucleotide through the nanopore.
11. A kit comprising:
(i) a device comprising a structure having a nanopore aperture that separates two independent adjacent pools of solution; and
(ii) a helicase that is capable of moving a polynucleotide with respect to the nanopore aperture a rate of less than 250 Hz.
12. The kit of claim 11 , wherein the nanopore aperture is solid-state.
13. The kit of claim 11 , wherein the nanopore aperture comprises a biological nanopore.
14. The kit of claim 11 , wherein the helicase is selected from the group consisting of E - coli bacteriophage T7 gp4 and T4 gp41 gene proteins, and the E. coli proteins DnaB, RuvB, and rho.
15. The kit of claim 11 , wherein the nanopore device further comprises electrodes at or near the nanopore aperture.
16. The kit of claim 11 , wherein the nanopore device further comprises electrodes placed within the adjacent pools.
17. The kit of claim 11 , further comprising a biological nanopore.
18. The kit of claim 11 , wherein the rate of the helicase is less than or equal to 200 Hz.
19. The kit of claim 11 , wherein the rate of the helicase is less than or equal to 150 Hz.
20. The kit of claim 11 , wherein the helicase is capable of moving the polynucleotide through the nanopore.
21. A method for analyzing a polynucleotide, said method comprising:
(i) providing a structure having a nanopore aperture that separates two independent adjacent pools of solution, wherein a helicase is located in at least one of the pools;
(ii) placing the polynucleotide in one of the pools and allowing the helicase to move the polynucleotide with respect to the nanopore aperture at a rate of less than 250 Hz; and
(iii) measuring transport properties of the polynucleotide as it moves with respect to the nanopore, thereby analyzing the polynucleotide.
22. The method of claim 21 , wherein the nanopore aperture is solid-state.
23. The method of claim 21 , wherein the nanopore aperture comprises a biological nanopore.
24. The method of claim 21 , wherein the transport properties are the amplitude or duration of individual conductance or electron tunneling current changes across the nanopore aperture.
25. The method of claim 21 , wherein the helicase is selected from the group consisting of E - coli l bacteriophage T 7 gp4 and T4 gp41 gene proteins, and the E. coli proteins DnaB, RuvB, and rho.
26. The method of claim 21 , wherein the device further comprises electrodes that are at or near the nanopore aperture and that measure the transport properties of the polynucleotide.
27. The method of claim 21 , wherein the device further comprises electrodes that placed within the adjacent pools and that measure the transport properties of the polynucleotide.
28. The method of claim 21 , wherein the rate of the helicase is less than or equal to 200 Hz.
29. The method of claim 21 , wherein the rate of the helicase is less than or equal to 150 Hz.
30. The method of claim 21 , further comprising determining the sequence of the polynucleotide from the measured transport properties.
31. The method of claim 21 , wherein, in step (ii), the helicase moves the polynucleotide through the nanopore.