Nanonozzle device arrays: their preparation and use for macromolecular analysis

• US 9,845,238 B2
• Filed: 05/14/2015
• Issued: 12/19/2017
• Est. Priority Date: 07/19/2006
• Status: Active Grant

• Current Assignee: BioNano Genomics, Inc.
• Original Assignee: BioNano Genomics, Inc.
• Inventors: Cao, Han, Deshpande, Parikshit A., Austin, Michael David, Boyce-Jacino, Michael
• Primary Examiner(s): Bhat, Narayan
• Application Number:
  US14/712,816
• Publication Number:
  US 20150323518A1
• Time in Patent Office: 950 Days
• CPC Class Codes: B01L 2200/0663 Stretching or orienting elongated molecules or particles
• CPC Class Codes: B01L 2200/12 Specific details about manufacturing devices
• CPC Class Codes: B01L 2300/0645 Electrodes
• CPC Class Codes: B01L 2300/0816 Cards, e.g. flat sample carriers usually with flow in two horizontal directions
• CPC Class Codes: B01L 2300/0822 Slides
• CPC Class Codes: B01L 2300/0896 Nanoscaled
• CPC Class Codes: B01L 2400/0415 electrical forces, e.g. electrokinetic
• CPC Class Codes: B01L 2400/0487 fluid pressure, pneumatics
• CPC Class Codes: B01L 3/502761 specially adapted for handling suspended solids or molecules independently from the bulk fluid flow, e.g. for trapping or sorting beads, for physically stretching molecules investigating characteristics of particles G01N15/00
• CPC Class Codes: B82Y 30/00 Nanotechnology for materials or surface science, e.g. nanocomposites
• CPC Class Codes: C12Q 1/6869 Methods for sequencing
• CPC Class Codes: C12Q 2565/631 being a biochannel or pore
• CPC Class Codes: G01N 2035/1039 Micropipettes, e.g. microcapillary tubes
• CPC Class Codes: G01N 27/447 using electrophoresis
• 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
• CPC Class Codes: Y10T 436/143333 Saccharide [e.g., DNA, etc.]
• CPC Class Codes: Y10T 436/2575 Volumetric liquid transfer

Abstract

Constricted nanochannel devices suitable for use in analysis of macromolecular structure, including DNA sequencing, are disclosed. Also disclosed are methods for fabricating such devices and for analyzing macromolecules using such devices.

Claims

1. A method for characterizing one or more features of a macromolecule, comprising:

linearizing the macromolecule residing at least in part within a nanochannel, said macromolecule comprising a label,

at least a portion of the nanochannel being capable of physically constraining at least a portion of the macromolecule so as to maintain in linear form that portion of the macromolecule, and

wherein a first effective inner diameter of the nanochannel is between about 10 nm and about 500 nm, the nanochannel comprising at least one constriction that is configured to locally reduce the effective inner diameter to a second effective inner diameter of about 0.5 nm to about 100 nm, whereby the second effective inner diameter is less than the first effective inner diameter; and

transporting at least a portion of the macromolecule within at least the portion of the nanochannel such that at least the portion of the macromolecule passes through the constriction;

monitoring at least one signal arising in connection with the label of the macromolecule passing through the constriction,

wherein the at least one signal characterizes one or more features of the macromolecule.

2. The method of claim 1, wherein the macromolecule is selected from the group consisting of:

a polynucleotide, a polynucleoside, a polymer, a copolymer, a dendrimer, a surfactant, a lipid, a carbohydrate, a polypeptide, a protein, and any combination thereof.

3. The method of claim 1, wherein the macromolecule resides within a fluid.

4. The method of claim 1, wherein the transporting comprises exposing the macromolecule to a gradient.

5. The method of claim 4, wherein the gradient is selected from the group consisting of:

an electroosmotic field, an electrophoretic field, a magnetic field, an electric field, an electromagnetic field, a flow field, a radioactive field, a mechanical force, an electroosmotic force, an electrophoretic force, an electrokinetic force, a temperature gradient, a pressure gradient, a surface property gradient, a capillary flow, and any combination thereof.

6. The method of claim 1, wherein the at least one signal is selected from the group consisting of:

a visual signal, an infrared signal, an ultraviolet signal, a radioactive signal, a magnetic signal, an electrical signal, an electromagnetic signal, or any combination thereof.

7. The method of claim 1, wherein the macromolecule comprises two or more labels.

8. The method of claim 1, wherein a feature of the macromolecule comprises the location of one or more labels of the macromolecule.

9. The method of claim 8, further comprising the detection of sites selected from the group consisting of:

binding sites between a drug and the macromolecule, macromolecule-drug complexes, DNA repairing sites, DNA binding sites, DNA cleaving sites, SiRNA binding sites, anti-sense binding sites, transcription factor binding sites, regulatory factor binding sites, restriction enzyme binding sites, restriction enzyme cleaving sites, and any combination thereof.

10. The method of claim 1, wherein the label is selected from the group consisting of:

an electron spin resonance molecule, a fluorescent molecule, a chemiluminescent molecule, a radioisotope, and enzyme substrate, a biotin molecule, an avidin molecule, an electrical charged transferring molecule, a semiconductor nanocrystal, a semiconductor nanoparticle, a colloid gold nanocrystal, a ligand, a microbead, a magnetic bead, a paramagnetic particle, a quantum dot, a chromogenic substrate, an affinity molecule, a protein, a peptide, a nucleic acid, a carbohydrate, an antigen, a hapten, an antibody, an antibody fragment, a lipid, a polymer, an electrically charged particle, modified nucleotide, and any combination thereof.

11. The method of claim 1, wherein a feature of the macromolecule is selected from the group consisting of:

the size of the macromolecule, the molecular composition of the macromolecule, the molecular sequence of the macromolecule, an electrical property of one or more molecules of the macromolecule, a chemical property of one or more molecules of the macromolecule, a radioactive property of one or more molecules of the macromolecule, a magnetic property of one or more molecules of the macromolecule, and any combination thereof.

12. The method of claim 11, wherein the molecular composition of the macromolecule is selected from the group consisting of:

the position of one or more molecules of the macromolecule, DNA polymorphisms, DNA copy number polymorphisms, amplifications within DNA, deletions within DNA, translocations within DNA, inversions of particular loci within DNA, the location of a methyl group within the macromolecule, and any combination thereof.

13. The method of claim 1, wherein a ratio of the second effective inner diameter to the first effective inner diameter of the nanochannel is at most 40%.

14. A method for transporting a macromolecule, comprising:

providing at least a first and a second fluid reservoir;

providing an at least partially linearized macromolecule,

at least a portion of the partially linearized macromolecule residing in a nanochannel, wherein a first effective inner diameter of the nanochannel is between about 10 nm and about 500 nm,

the nanochannel placing the at least two reservoirs in fluid communication with one another,

the nanochannel comprising a constriction in or at a terminal end of the nanochannel, said constriction configured to locally reduce the first effective inner diameter of the nanochannel to be about 0.5 nm to about 100 nm, whereby the second effective inner diameter is less than the first effective inner diameter; and

applying a gradient to the partially linearized macromolecule,

the gradient giving rise to at least the portion of the partially linearized macromolecule being transported within at least a portion of the nanochannel and to the second fluid reservoir; and

detecting signals from the second reservoir, said signals arising from the partially linearized macromolecule that has been transported to the second reservoir.

15. The method of claim 14, wherein the nanochannel comprises a length greater than about 100 nm.

16. The method of claim 14, wherein the nanochannel comprises a length greater than about the length of the partially linearized macromolecule.

17. The method of claim 14, wherein the gradient is selected from the group consisting of:

an electroosmotic field, an electrophoretic field, capillary flow, a magnetic field, an electric field, a radioactive field, a mechanical force, an electroosmotic force, an electrophoretic force, an electrokinetic force, a temperature gradient, a pressure gradient, a surface property gradient, a capillary flow, and any combination thereof.

18. A method for linearizing a macromolecule, comprising:

placing the macromolecule in a nanochannel, wherein the nanochannel comprises a constriction in or at a terminal end of the nanochannel,

at least a portion of the nanochannel being capable of physically constraining at least a portion of the macromolecule so as to maintain in linear form that portion of the macromolecule,

wherein a first effective inner diameter of the nanochannel is between about 10 nm and about 500 nm, and wherein the constriction is configured to locally reduce the first effective inner diameter of the nanochannel to a second effective inner diameter that is less than about 40% of the first effective inner diameter, wherein the second effective inner diameter is less than the first effective inner diameter, whereby the macromolecule is linearized.

19. The method of claim 18, further comprising applying a gradient to the macromolecule, such that at least a portion of the macromolecule passes, linearly, through the nanochannel constriction.

20. The method of claim 19, wherein the gradient is selected from the group consisting of:

an electroosmotic field, an electrophoretic field, capillary flow, a magnetic field, an electric field, a radioactive field, a mechanical force, an electroosmotic force, an electrophoretic force, an electrokinetic force, a temperature gradient, a pressure gradient, a surface property gradient, a capillary flow, and any combination thereof.

21. The method of claim 18, wherein the nanochannel places two or more fluid reservoirs in fluid communication with one another.

22. The method of claim 18, wherein the first effective inner diameter of the nanochannel is less than about two times the radius of gyration of the linear conformation of the macromolecule.