DEVICES, SYSTEMS, AND METHODS FOR AMPLIFYING NUCLEIC ACIDS

• US 20100304446A1
• Filed: 03/26/2010
• Published: 12/02/2010
• Est. Priority Date: 02/07/2006
• Status: Abandoned Application

• Current Assignee: Stokes Bio Ltd. (Perspecta, Inc.), Life Technologies Corporation (Thermo Fisher Scientific Incorporated)
• Original Assignee: Stokes Bio Ltd. (Perspecta, Inc.)
• Inventors: Davies, Mark, Dalton, Tara
• Application Number:
  US12/732,596
• Publication Number:
  US 20100304446A1
• Time in Patent Office: Days
• US Class Current: 435/91.21
• CPC Class Codes: B01L 2200/0673 Handling of plugs of fluid surrounded by immiscible fluid
• CPC Class Codes: B01L 2400/0457 passive flow or gravitation
• CPC Class Codes: B01L 2400/0487 fluid pressure, pneumatics
• CPC Class Codes: B01L 3/502784 specially adapted for droplet or plug flow, e.g. digital microfluidics automatic analysis using a stream of discrete samples in a tube system G01N35/08
• CPC Class Codes: B01L 7/525 with physical movement of samples between temperature zones
• CPC Class Codes: C12Q 1/6851 Quantitative amplification
• CPC Class Codes: C12Q 2561/113 Real time assay
• CPC Class Codes: C12Q 2565/629 being a microfluidic device

Abstract

The present invention generally relates to a devices, systems, and methods for amplifying nucleic acids in flowing droplets. In certain embodiments, the invention provides a device for amplifying nucleic acids including at least one channel through which sample droplets including nucleic acids flow, in which the nucleic acids in the droplets are optically detectable while the droplets are flowing through the channel, and a plurality of temperature zones in thermal contact with the channel, in which the zones are located at different locations along the channel and the zones are separated from each other.

Claims

1. A device for amplifying nucleic acids, the device comprising:

at least one first channel through which sample droplets comprising nucleic acids flow, wherein the nucleic acids in the droplets are optically detectable while the droplets are flowing through the first channel; and

a plurality of temperature zones in thermal contact with the first channel, wherein the zones are located at different locations along the first channel and the zones are separated from each other.

2. The device according to claim 1, wherein a first temperature zone provides a temperature sufficient to result in denaturation of double stranded nucleic acids to produce single stranded nucleic acids, a second temperature zone provides a temperature sufficient to result in hybridization of primers to the single stranded nucleic acids, and a third temperature zone provides a temperature sufficient to result in amplification of single stranded nucleic acids to produce double stranded nucleic acids.

3. The device according to claim 2, wherein the plurality of temperature zones are arranged as repeating first, second, and third zones.

4. The device according to claim 1, further comprising a second channel, wherein the second channel is open at a top and the first channel lies within the second channel.

5. The device according to claim 1, wherein the device is coupled to an excitation system.

6. The device according to claim 5, wherein the device is coupled to an optical detection device.

7. The system according to claim 6, wherein the optical detection device is a spectrograph.

8. The device according to claim 1, wherein the device is fluidically coupled to a liquid bridge system.

9. The device according to claim 1, wherein the droplets are wrapped in an immiscible fluid.

10. The device according to claim 1, wherein air gaps isolate the temperature zones from each other.

11. The device according to claim 1, wherein the nucleic acid is DNA or RNA.

12. A system for amplifying nucleic acids, the system comprising:

a sample acquisition stage;

a device for mixing sample droplets to form mixed droplets comprising nucleic acids wrapped in an immiscible carrier fluid; and

a device for amplifying the nucleic acids comprising at least one first channel through which the sample droplets flow, wherein the nucleic acids in the droplets are optically detectable while the droplets are flowing through the first channel; and

a plurality of temperature zones in thermal contact with the first channel, wherein the zones are located at different locations along the first channel and the zones are separated from each other.

13. The system according to claim 12, further comprising an excitation system.

14. The system according to claim 13, further comprising an optical detection device.

15. The system according to claim 14, wherein the optical detection device is a spectrograph

16. The system according to claim 12, wherein the device for amplifying the nucleic acids further comprises a second channel, wherein the second channel is open at a top and the first channel lies within the second channel.

17. The system according to claim 12, wherein a first temperature zone provides a temperature sufficient to result in denaturation of double stranded nucleic acids and a second temperature zone provides a temperature sufficient to result in regeneration of double stranded nucleic acids.

18. The system according to claim 17, wherein the plurality of temperature zones are arranged as alternating first and second zones.

19. The system according to claim 12, wherein the droplet forming device is a liquid bridge.

20. The system according to claim 12, wherein air gaps isolate the temperature zones from each other.

21. A device for applying a temperature profile to a sample molecule, the device comprising:

at least one first channel through which sample droplets comprising sample molecules flow, wherein the molecules in the droplets are optically detectable while the droplets are flowing through the first channel; and

a plurality of temperature zones in thermal contact with the first channel, wherein the zones are located at different locations along the first channel and the zones are separated from each other.

22. A method for performing a quantitative polymerase chain reaction, the method comprising the steps of:

a) flowing sample droplets comprising labeled nucleic acids through at least one first channel, wherein the labeled nucleic acids in the droplets are optically detectable while the droplets are flowing through the first channel;

b) denaturing double stranded nucleic acids in the flowing sample droplets to produce single stranded nucleic acids;

c) hybridizing primers to the single stranded nucleic acids in the flowing sample droplets;

d) simultaneously amplifying the nucleic acids and detecting the amplified nucleic acids in the flowing droplets; and

e) repeating steps (b) through (d) at least once.

23. The method according to claim 22, wherein the nucleic acid is DNA or RNA.

24. The method according to claim 22, wherein the droplets are wrapped in an immiscible fluid.

25. The method according to claim 22, wherein prior to step (a), the method further comprises forming sample droplets using a liquid bridge system.

26. The method according to claim 22, wherein the first channel lies within a second channel that is open at a top.