Nanofluidic sorting system for gene synthesis and pcr reaction products

Patent metadata

Field	Value
Publication number	US-10065154-B2
Application number	US-201314433471-A
Country	US
Kind code	B2
Filing date	Oct 4, 2013
Priority date	Oct 5, 2012
Publication date	Sep 4, 2018
Grant date	Sep 4, 2018

Abstract

Devices and methods integrate nanopore and microfluidic technologies for recording molecular characteristics of individual molecules such as, for example, biomolecules. Devices comprise a first substrate comprising a microchannel, a second substrate comprising a microchannel, the second substrate positioned below the first substrate, and a membrane having a thickness of about 0.3 nm to about 1 nm and comprising at least one nanopore, the membrane positioned between the first substrate and the second substrate, wherein a single nanopore of the membrane is constructed and arranged for electrical and fluid communication between the microchannel of the first substrate and the microchannel of the second substrate. To mitigate the effect of errors that occur during de novo DNA synthesis, longer DNA molecules are typically synthesized from shorter oligonucleotides by polymerase construction and amplification (PCA), or by other methods.

First claim

What is claimed is:

1. A device comprising
a first substrate comprising at least one microchannel,
a second substrate comprising at least one microchannel, the second substrate positioned below the first substrate, and
a membrane having a thickness of about 0.3 nm to about 1 μm and comprising at least one nanopore, the membrane positioned between the first substrate and the second substrate,
wherein a nanopore of the membrane is constructed and arranged for electrical and fluid communication at an intersection between a microchannel of the first substrate and a microchannel of the second substrate, wherein a substantially horizontal axis of a microchannel of the first substrate is positioned at an angle of about 10° to about 90° relative to a substantially horizontal axis of a microchannel of the second substrate.

2. The device of claim 1 , wherein the membrane has a thickness of about 0.3 nm to about 500 nm.

3. The device of claim 1 , wherein one side of the membrane has a surface area of about 10 μm×10 μm to about 10 mm×10 mm.

4. The device of claim 1 , wherein the membrane is a dielectric membrane.

5. The device of claim 4 , wherein the dielectric membrane is a silicon nitride (SiNx) dielectric membrane.

6. The device of claim 1 , wherein the membrane is coated with a semiconductor material.

7. The device of claim 6 , wherein the semiconductor material is at least one of alumina (Al 2 O 3 ), silicon dioxide (SiO 2 ), hafnium dioxide (HfO 2 ), titanium dioxide (TiO 2 ), graphene, hexagonal boron nitride (hBN), zinc oxide (ZnO), indium arsenide (InAs), bismuth selenide (BiSe), bismuth telride (BeTe 2 ), lead selenide (PbSe 2 ), nickel silicide (NiSi), tungsten diselenide (WSe 2 ), copper oxide (CuO), gallium nitride (GaN), molybdenum disulfide (MoS 2 ), niobium diselenide (NbSe 2 ), and Bi 2 Sr 2 CaCu 2 O.

8. The device of claim 1 , wherein a microchannel of the first and/or second substrate has an inlet at a first end and an outlet at a second end.

9. The device of claim 8 , further comprising a negative electrode at the inlet of a microchannel of the first substrate, and a positive electrode at the outlet of a microchannel of the second substrate.

10. The device of claim 1 , wherein the first and/or second substrate comprises 2 to 2000 microchannels.

11. The device of claim 1 , wherein a microchannel of the first and/or second substrate is a substantially linear microchannel or has a branched portion at one or more ends.

12. The device of claim 11 , wherein the branched portion comprises 2 to 20 microchannels.

13. The device of claim 1 , wherein a microchannel of the first and/or second substrate has a collection chamber at one or more ends.

14. The device of claim 1 , wherein the device comprises at least one access port.

15. The device of claim 1 , wherein the central portion of a microchannel of the first and/or second substrate has a width of about 100 nm to about 1 mm.

16. The device of claim 1 , wherein the at least one nanopore of the membrane has a diameter of about 0.2 nm to about 1 μm and a length of about 0.3 nm to about 1 μm.

17. The device of claim 1 , wherein the at least one nanopore of the membrane has a diameter that is constructed and arranged for translocation from one microchannel to another microchannel of a nucleic acid molecule that is 20 nucleotides to 10 6 nucleotides in length.

18. The device of claim 17 , wherein the nucleic acid molecule is a single-stranded nucleic acid molecule or a double-stranded nucleic acid molecule.

19. The device of claim 1 , wherein the membrane comprises 2 to 10000 nanopores.

20. The device of claim 1 , wherein each substrate comprises a polymer or a non-polymer.

21. The device of claim 20 , wherein each substrate comprises a polymer that comprises silicone, polydimethylsiloxane (PDMS), polycarbonate, poly(methyl methacrylate), zeonax, cyclic olefin polymer (COP), polyester toner (PeT) and cellulose.

22. The device of claim 20 , wherein each substrate comprises a non-polymer that comprises glass, silica, silicon, nitride, paper, gallium arsenide or germanium.

23. The device of claim 1 , wherein the first and/or second substrate comprises at least one surface modification selected from the group consisting of a crosslinking agent, a silane group, an adhesive coating and plasma.

24. The device of claim 1 , wherein one side of the first and/or second substrate has a surface area of about 50 μm 2 to about 100 mm 2 .

25. The device of claim 1 , wherein the surface area of one side of the first substrate is about equal with the surface area of one side of the second substrate.

26. The device of claim 1 , wherein the membrane is covalently bonded to the first and/or second substrate.

27. The device of claim 1 , wherein the device is connected to a switch, amplifier, digital recorder, computer or a combination thereof.

28. An array comprising 2 to 10 5 devices of claim 1 .

29. A method, comprising adding a plurality of molecules to a microchannel of the first or second substrate of the device of claim 1 ;
applying an ionic current across the membrane, thereby providing for translocation of a molecule of the plurality of molecules from a microchannel of one substrate to a microchannel of another substrate through a nanopore of the membrane.

30. A device comprising
a first substrate comprising at least one microchannel,
a second substrate comprising at least one microchannel, the second substrate positioned below the first substrate,
a membrane having a thickness of about 0.3 nm to about 1 μm and comprising at least one nanopore, the membrane positioned between the first substrate and the second substrate,
wherein a nanopore of the membrane is constructed and arranged for electrical and fluid communication at an intersection between a microchannel of the first substrate and a microchannel of the second substrate, and
wherein the device further comprises an additional substrate positioned above or below the first and/or second substrate, the additional substrate comprising
at least one microchannel;
an additional membrane having a thickness of about 0.3 nm to about 1 μm and comprising at least one additional nanopore;
an additional membrane having a thickness of about 0.3 nm to about 1 μm disposed between an additional substrate and the first and/or second substrate, wherein the additional membrane comprises at least one additional nanopore; or
one or more valves.

31. A method comprising
providing a first substrate comprising at least one microchannel, wherein a membrane having a thickness of about 0.3 nm to about 1 μm and comprising at least one nanopore is disposed on a surface of the first substrate such that the at least one nanopore of the membrane contacts the microchannel of the first substrate;
providing a second substrate comprising at least one microchannel; and
contacting the membrane with the second substrate, wherein a nanopore of the membrane provides electrical and fluid communication between the microchannel of the first substrate and the microchannel of the second substrate, wherein a substantially horizontal axis of a microchannel of the first substrate is positioned at an angle of about 10° to about 90° relative to a substantially horizontal axis of a microchannel of the second substrate.

32. A

Assignees

• Massachusetts Inst Technology

Inventors

• Aguilar Carlos Andres
• Jain Tarun Kumar
• Karnik Rohit N
• Carr Peter A

Classifications

• G01N27/44756

  Apparatus specially adapted therefor · CPC title

• B32B37/26

  with at least one layer which influences the bonding during the lamination process, e.g. release layers or pressure equalising layers · CPC title

• G01N33/48721

  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 title

• G01N15/12

  by observing changes in resistance or impedance across apertures when traversed by individual particles, e.g. by using the Coulter principle · CPC title

• B01D63/005

  Microfluidic devices (Microfluidic devices comprising semi-permeable hollow fibre membranes B01D63/028; Microfluidic devices comprising semi-permeable flat membranes B01D63/088) · CPC title