Nanopore and Carbon Nanotube Based Dna Sequencer and A Serial Recognition Elements

• US 20130186757A1
• Filed: 10/06/2009
• Published: 07/25/2013
• Est. Priority Date: 10/06/2008
• Status: Active Grant

• Current Assignee: Arizona Board of Regents (University of Arizona)
• Original Assignee: Arizona Board of Regents (University of Arizona)
• Inventors: Reinhart, Kevin, Lindsay, Stuart, Zhang, Peiming
• Granted Patent: US 8,968,540 B2
• Time in Patent Office: Days
• US Class Current: 204/452
• CPC Class Codes: C07D 235/24 with hetero atoms or with carbon atoms having three bonds to hetero atoms with at the most one bond to halogen, e.g. ester or nitrile radicals, directly attached in position 2
• CPC Class Codes: C12Q 1/6869 Methods for sequencing
• CPC Class Codes: C12Q 2565/631 being a biochannel or pore
• CPC Class Codes: G01N 27/447 using electrophoresis
• CPC Class Codes: G01N 27/44791 Microapparatus sample containers with integrated microfluidic structures B01L3/5027
• CPC Class Codes: G01N 33/48721 Investigating individual macromolecules, e.g. by translocation through nanopores Coulter counters in general G01N15/12; fabrication methods for nanoscale apertures B81B1/00; sequencing of nucleic acids C12Q1/68

Abstract

The present invention is directed to systems, devices and methods for identifying biopolymers, such as strands of DNA, as they pass through a constriction such as a carbon nanotube nanopore. More particularly, the invention is directed to such systems, devices and methods in which a newly translocated portion of the biopolymer forms a temporary electrical circuit between the nanotube nanopore and a second electrode, which may also be a nanotube. Further, the invention is directed to such systems, devices and methods in which the constriction is provided with a functionalized unit which, together with a newly translocated portion of the biopolymer, forms a temporary electrical circuit that can be used to characterize that portion of the biopolymer.

Claims

1. A molecular recognition chip configured to identify at least one portion of a target molecule, the chip comprising:

a) a substrate;

b) a first nanotube passing through at least a portion of the substrate, the first nanotube provided with a nanopore and configured and dimensioned to permit the translocation of a target molecule therethrough;

c) a second nanotube supported by the substrate and disposed in a fixed relationship with respect to said first nanotube so as to form a first gap with the first nanotube; and

d) an electrical circuit configured to detect an electrical current between the first and second nanotubes, upon passage of the target molecule past said gap;

wherein the first nanotube comprises a first recognition element connected to an end thereof, the second nanotube comprises a second recognition element connected to a first end thereof, and wherein each of the first and second recognition elements further comprise a universal base reader.

2. The chip of claim 1 wherein the universal base reader is capable of recognizing different DNA bases and is also capable of forming an additional set of hydrogen bonds with a complex of a DNA base and the universal base reader.

3. The chip of claim 2 wherein the base reader comprises 4-(mercaptomethyl)-1H-imidazole-2-carboxamide.

4. The chip of claim 2 wherein the base reader comprises 5-(2-aminoethyl)-1H-imidazole-2-carboxamide.

5. The chip of claim 2 wherein the base reader comprises thiobenzoic acid.

7. The device of claim 5, wherein the universal base readers are selected, independently, from the group consisting of:

8. The device of claim 5 wherein the universal base readers comprise 4-(mercaptomethyl)-1H-imidazole-2-carboxamide.

9. The device of claim 5 wherein the universal base readers comprise 5-(2-aminoethyl)-1H-imidazole-2-carboxamide.

10. The device of claim 5 wherein the universal base reader comprises thiobenzoic acid.

11. The device of claim 5 wherein the universal base readers comprise a conjugate of a planar π

system with a functional group, amine, amide, nitro, carboxylic acid, and halogen, connected to a molecular linker or a conjugate of the functional group with a molecular linker.

12. The device of claim 9 wherein the molecular linker is functionalized with an anchoring group selected from the group consisting of amine, thiol, carboxylic acid, halogen, selenium, and hydroxyl.

13. A method of sequencing DNA as the DNA molecule translocates through a constriction, the method comprising:

e) providing an apparatus comprising a device in accordance with claim 5, the device being located in the apparatus such that a first chamber is located on a first side of the device and a second chamber is located on a second side of the device;

f) introducing the DNA molecule into the second chamber;

g) electrophoresing the DNA molecule so that it translocates through the constriction; and

h) detecting an electrical current passing through the first electrode, a first universal base reader, a nucleotide base of the DNA molecule, a second universal base reader, and the second electrode.

14. The method of claim 12 wherein the first and second universal base reader are the same.

15. The method of claim 13 wherein the universal base reader comprises thiobenzoic acid.

6. A device for obtaining an electrical signal characteristic of a DNA base pair comprising a first universal base reader attached to a first electrode and a second universal base reader attached to a second electrode, wherein the universal base readers are capable of recognizing different DNA bases and are also capable of forming an additional set of hydrogen bonds with a complex of a DNA base and the universal base reader.

16. A composition comprising a universal base reader wherein the reader is selected from the group consisting of 4-(mercaptomethyl)-1H-imidazole-2-carboxamide and 5-(2-aminoethyl)-1H-imidazole-2-carboxamide.

17. A method of synthesizing 5-(2-aminoethyl)-1H-imidazole-2-carboxamide, the method as shown in FIG. 55.