Nucleic acid sample preparation

• US 9,827,565 B2
• Filed: 10/13/2016
• Issued: 11/28/2017
• Est. Priority Date: 04/16/2012
• Status: Active Grant

• Current Assignee: Biological Dynamics, Inc.
• Original Assignee: Biological Dynamics, Inc.
• Inventors: Krishnan, Rajaram, Charlot, David, Tu, Eugene, McCanna, James, Kumosa, Lucas, Swanson, Paul, Turner, Robert, Yang, Kai, Dobrovolskaya, Irina, Liu, David
• Primary Examiner(s): Martinell, James
• Application Number:
  US15/293,062
• Publication Number:
  US 20170072395A1
• Time in Patent Office: 411 Days
• Field of Search: None
• CPC Class Codes: B01D 57/02 by electrophoresis treatment of water, waste water, sewage or sludge by electrophoresis C02F1/469; electrophoretic production of compounds or non-metals C25B7/00; investigating or analysing materials by using electrophoresis G01N27/26
• CPC Class Codes: B01L 2300/0636 Integrated biosensor, microarrays
• CPC Class Codes: B01L 2300/0645 Electrodes
• CPC Class Codes: B01L 2400/0424 Dielectrophoretic forces
• CPC Class Codes: B01L 3/502715 characterised by interfacing components, e.g. fluidic, electrical, optical or mechanical interfaces
• CPC Class Codes: B01L 7/52 with provision for submitting samples to a predetermined sequence of different temperatures, e.g. for treating nucleic acid samples amplification or hybridisation processes per se C12Q1/68; controlling sequential reactions for synthesis B01J19/0046
• CPC Class Codes: B03C 2201/26 for use in medical applications
• CPC Class Codes: B03C 5/005 Dielectrophoresis, i.e. dielectric particles migrating towards the region of highest field strength B03C5/02 takes precedence; electrophoresis B01D57/02
• CPC Class Codes: B03C 5/026 using open-gradient differential dielectric separation, i.e. using electrodes of special shapes for non-uniform field creation, e.g. Fluid Integrated Circuit [FIC]
• CPC Class Codes: B03C 7/023 Non-uniform field separators
• CPC Class Codes: C12N 15/1003 Extracting or separating nucleic acids from biological samples, e.g. pure separation or isolation methods; Conditions, buffers or apparatuses therefor
• CPC Class Codes: C12N 15/101 by chromatography, e.g. electrophoresis, ion-exchange, reverse phase
• CPC Class Codes: C12Q 1/6806 Preparing nucleic acids for analysis, e.g. for polymerase chain reaction [PCR] assay C12Q1/6804 takes precedence
• CPC Class Codes: C12Q 2523/307 Denaturation or renaturation by electric current/voltage
• CPC Class Codes: G01N 27/447 using electrophoresis
• CPC Class Codes: G01N 27/44704 Details; Accessories
• CPC Class Codes: G01S 11/12 using electromagnetic waves other than radio waves
• CPC Class Codes: G01S 5/163 Determination of attitude using inertial means G01C9/00; control of attitude G05D1/49
• CPC Class Codes: G06F 3/0346 with detection of the device orientation or free movement in a 3D space, e.g. 3D mice, 6-DOF [six degrees of freedom] pointers using gyroscopes, accelerometers or tilt-sensors

Abstract

The present invention includes methods, devices and systems for isolating a nucleic acid from a fluid comprising cells. In various aspects, the methods, devices and systems may allow for a rapid procedure that requires a minimal amount of material and/or results in high purity nucleic acid isolated from complex fluids such as blood or environmental samples.

Claims

1. A device for isolating DNA from a fluid comprising cells, the device comprising:

a. a first array of electrodes capable of establishing a first AC electrokinetic field region, wherein the first AC electrokinetic field region is a first dielectrophoretic low field region capable of concentrating and lysing a plurality of cells in a conductivity of fluid of 100 mS/m or more;

b. a second array of electrodes capable of establishing a second AC electrokinetic field, wherein the second AC eletrokinetic field is a dieletrophoretic high field region capable of concentrating and isolating nucleic acid in a conductivity of fluid of 100 mS/m or more;

c. a reservoir comprising a fluid and a device for flushing residual and lysed cellular material from the first array of electrodes; and

d. a module capable of performing sequencing.

2. The device of claim 1, wherein the first and second array of electrodes are configured to be selectively energized to establish a dielectrophoretic high field and dielectrophoretic low field regions.

3. The device of claim 1, wherein the first array of electrodes and the second array of electrodes is in a wavy or nonlinear line configuration, wherein the configuration comprises a repeating unit comprising the shape of a pair of dots connected by a linker, wherein the dots and linker define the boundaries of the electrode, wherein the linker tapers inward towards or at the midpoint between the pair of dots, wherein the diameters of the dots are the widest points along the length of the repeating unit, wherein the edge to edge distance between a parallel set of repeating units is equidistant, or roughly equidistant.

4. The device of claim 1, wherein nucleic acid is isolated and sequencing is performed in a single chamber.

5. The device of claim 1, wherein nucleic acid is isolated and sequencing is performed in multiple regions of a single chamber.

6. The device of claim 1, further comprising using at least one of an elution tube, a chamber and a reservoir to perform sequencing.

7. The device of claim 1, wherein sequencing of the nucleic acid is performed in a serpentine microchannel comprising a plurality of temperature zones.

8. The device of claim 1, wherein sequencing is performed in aqueous droplets entrapped in immiscible fluids.

9. The device of claim 1, wherein the device comprises a module capable of thermocycling and amplifying nucleic acids.

10. The device of claim 1, wherein the device comprises a surface contacting or proximal to the electrodes, wherein the surface is functionalized with biological ligands that are capable of selectively capturing biomolecules.

11. The device of any of claim 1, wherein the array of electrodes is coated with a hydrogel.

12. The device of claim 11, wherein the hydrogel comprises two or more layers of a synthetic polymer.

13. The device of claim 11, wherein the hydrogel is spin-coated onto the electrodes.

14. The device of claim 11, wherein the hydrogel has a viscosity between about 0.5 cP to about 5 cP prior to spin-coating.

15. The device of any claim 11, wherein the hydrogel has a thickness between about 0.01 microns and 1 micron.

16. The device of any of claim 11, wherein the hydrogel has a conductivity between about 0.1 S/m to about 1.0 S/m.

17. The device of claim 1, wherein the module capable of performing sequencing is a module capable of performing Sanger sequencing.

18. The device of claim 17, wherein the module capable of performing Sanger sequencing comprises a module capable of capillary electrophoresis, a module capable of multi-color fluorescence detection, or a combination thereof.

19. The device of claim 1, wherein the module capable of performing sequencing is a module capable of performing next generation sequencing.

20. The device of claim 19, wherein the module capable of performing next-generation sequencing is capable of performing pyrosequencing, ion semiconductor sequencing, polony sequencing, sequencing by ligation, DNA nanoball sequencing, or single molecule sequencing.