Methods for fast nucleic acid amplification

Patent metadata

Field	Value
Publication number	US-10144960-B2
Application number	US-201815890843-A
Country	US
Kind code	B2
Filing date	Feb 7, 2018
Priority date	May 24, 2012
Publication date	Dec 4, 2018
Grant date	Dec 4, 2018

Abstract

Devices are provided for performing polymerase chain reaction (PCR).

First claim

The invention claimed is:

1. A device for performing PCR comprising;
a sample container for holding a PCR sample, wherein the sample container comprises
a target nucleic acid,
dNTPs,
a polymerase provided at a concentration of at least 0.5 μM, and
a pair of primers configured for amplifying the target nucleic acid, wherein the primers are each provided at a concentration of at least 2 μM;

means for heating the sample;
means for cooling the sample; and
control means for repeatedly subjecting the sample container to the means for heating the sample and the means for cooling the sample to thermal cycle the sample at a ramp rate of at least 200° C./s.

2. The device of claim 1 , further comprising means for optically exciting the sample in the monitoring position to cause the sample to fluoresce; and means for detecting the fluorescence of the excited sample during amplification.

3. The device of claim 2 , wherein the ramp rate is at least 300° C./s.

4. The device of claim 3 , wherein the ramp rate is at least 400° C./s.

5. The device of claim 1 , wherein the means for heating is a hot water bath and the means for cooling is a cool water bath.

6. The device of claim 1 , wherein the device is a microfluidics system.

7. A device for performing PCR comprising;
a sample container for holding a PCR sample, wherein the sample container comprises
a target nucleic acid,
dNTPs,
a polymerase provided at a concentration of at least 0.5 μM, and
a pair of primers configured for amplifying the target nucleic acid, wherein the primers are each provided at a concentration of at least 2 μM;

a heat transfer device for changing temperature of the sample in the sample container; and
a temperature controller for controlling the heat transfer device to repeatedly heat and cool the sample at a ramp rate of at least 200° C./s.

8. The device of claim 7 , wherein
the heat transfer device comprises a plurality of water baths, and
the temperature controller is a motor attached to an arm that moves the sample container between the water baths.

9. The device of claim 7 , wherein
the heat transfer device comprises a plurality of temperature zones,
the sample container defines a path in each of the temperature zones, and
the temperature controller controls flow of the sample to each of the temperature zones.

10. The device of claim 7 , wherein
the heat transfer device comprises heated air.

11. The device of claim 7 , wherein
the heat transfer device includes Joule heating.

12. The device of claim 7 , wherein the heat transfer device is a heat block.

13. The device of claim 7 , wherein the sample container further comprises a dye, and the device further comprises an optics block configured to detect emissions from the dye.

14. The device of claim 13 , wherein the optics block detects emissions from the dye in real-time.

15. The device of claim 7 , wherein the temperature controller is triggered by sample temperature.

16. The device of claim 7 , wherein the temperature controller is programmed to hold the sample at a temperature for a specific duration of time.

17. The device of claim 16 , wherein the specific duration of time is less than one second.

18. The device of claim 7 , wherein the device is controlled by a CPU.

19. The device of claim 7 , wherein transfer between temperature zones is achieved in less than 100 msec.

20. A device for performing PCR comprising;
a sample container for holding a PCR sample, wherein the sample container comprises eukaryotic genomic DNA and further comprises a polymerase provided at a concentration of at least 0.5 μM;
a heat transfer device for changing temperature of the sample in the sample container; and
a temperature controller for controlling the heat transfer device to repeatedly heat and cool the sample at a ramp rate of at least 200° C./s.

21. The device of claim 20 , wherein the sample container further comprises a pair of primers configured for amplifying the target nucleic acid, wherein the primers are each provided at a concentration of at least 2 μM.

22. The device of claim 20 , wherein the sample container further comprises primers for amplification of a target nucleic acid sequence of the eukaryotic genomic DNA, wherein the polymerase to primer ratio is 0.03 to 0.4 polymerase: total primer concentration.

Assignees

• Univ Utah Res Found

Inventors

• Wittwer Carl T
• Farrar Jared Steven

Classifications

• B01L7/5255

  by moving sample containers · CPC title

• B01L3/5027

  by integrated microfluidic structures, i.e. dimensions of channels and chambers are such that surface tension forces are important, e.g. lab-on-a-chip · CPC title

• B01L2300/0627

  Sensor or part of a sensor is integrated · CPC title

• C12Q1/686 Primary

  Polymerase chain reaction [PCR] · CPC title

• B01L2300/1894

  Cooling means; Cryo cooling · CPC title