POLYMERASE CHAIN REACTON METHOD, POLYMERASE CHAIN REACTON DROPLET DEVICE, AND POLYMERASE CHAIN REACTON DROPLET DEVICE ARRAY

• US 20110159547A1
• Filed: 02/10/2010
• Published: 06/30/2011
• Est. Priority Date: 12/29/2009
• Status: Active Grant

• Current Assignee: National Applied Research Laboratories (Government of The Republic of China)
• Original Assignee: National Applied Research Laboratories (Government of The Republic of China)
• Inventors: TSENG, FAN-GANG, HU, YI-CHIUEM, YU, CHIH-SHENG
• Granted Patent: US 8,389,273 B2
• Time in Patent Office: Days
• US Class Current: 435/91.2
• CPC Class Codes: B01L 2200/0673 Handling of plugs of fluid surrounded by immiscible fluid
• CPC Class Codes: B01L 2200/142 Preventing evaporation
• CPC Class Codes: B01L 2300/0819 Microarrays; Biochips
• CPC Class Codes: B01L 2300/1827 using resistive heater
• CPC Class Codes: B01L 3/5088 confining liquids at a location by surface tension, e.g. virtual wells on plates, wires B01L3/50857 takes precedence
• 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

Abstract

The present invention discloses a polymerase chain reaction (PCR) method, a PCR droplet device and a PCR droplet device array. The steps of the method comprise that a liquid comprising an analyzer is dropped on the heating coil disposed on the droplet device to form a droplet, then dropping a hydrophobic solution to prevent the droplet from evaporating. When an electric current or a voltage is supplied through at least one conducting wire to heat the heating coil, the inside of the droplet can generate buoyancy to drive the analyzer to move to the top of the inside of the droplet. Subsequently, the analyzer is moved to a periphery of the inside of the droplet so as to form a thermal cycle. Therefore the template is amplified by recycling the thermal cycle.

Claims

1. A polymerase chain reaction (PCR) method, comprising the following steps:

dropping a liquid comprising an analyzer onto a heating coil disposed on a droplet device so as to form a droplet, the analyzer comprising a template, a primer, a deoxyribonucleotide triphosphate (dNTP), and a polymerase;

dropping a hydrophobic solution onto a surface of the droplet to prevent the liquid from evaporating; and

applying an electric current or a voltage through at least one conducting wire disposed on the droplet device to heat the heating coil, such that buoyancy in an inside of the droplet is generated, thereby driving the analyzer to move to a top of the inside of the droplet, and subsequently to move to a periphery of the inside of the droplet so as to form a thermal cycle;

wherein, a temperature of a center of a bottom of the droplet is in a range of 90 to 100°

C. to denature the template, and the analyzer is moved to the top of the inside of the droplet by the buoyancy;

a temperature of the top of the inside of the droplet is in a range of 30 to 65°

C. to anneal the primer on a specific position of the template;

the analyzer is moved to a side of the bottom of the droplet by the circular shape of the droplet, and a temperature of the side of the bottom of the droplet is in a rage of 65 to 80°

C. so as to extend the specific position of the template; and

the analyzer is moved back to the center of the bottom of the droplet to form the thermal cycle, such that the template is amplified by recycling the thermal cycle.

2. The PCR method as claimed in claim 1, wherein the hydrophobic solution comprises mineral oil.

3. The PCR method as claimed in claim 1, wherein a metal electrical conductivity of the heating coil is higher than a metal electrical conductivity of the at least one conducting wire.

4. The PCR method as claimed in claim 3, wherein a material of the heating coil comprises silver, copper, gold, platinum, aluminum, iron, stannum, lead or a combination thereof.

5. The PCR method as claimed in claim 4, wherein a material of the at least one conducting wire comprises silver, copper, gold, platinum, aluminum, iron, stannum, lead or a combination thereof.

6. The PCR method as claimed in claim 1, wherein the template comprises deoxyribonucleic acid (DNA) or ribonucleic acid (RNA).

7. The PCR method as claimed in claim 1, wherein a shape of the heating coil comprises a circle, an angle, or an irregular shape.

8. The PCR method as claimed in claim 1, wherein the liquid comprises glycerol.

9. A polymerase chain reaction (PCR) droplet device, comprising:

a cooling plate;

a substrate disposed on the cooling plate;

a first containing area disposed on the substrate to contain a liquid comprising an analyzer;

a second containing area disposed on the substrate, and the second containing area surrounding the first containing area to contain a hydrophobic solution;

a heating coil disposed on a center of the first containing area;

at least one conducting wire disposed on the substrate and connected with the heating coil; and

a plurality of sensors disposed around the heating coil to detect temperature changes of the heating coil;

wherein, the analyzer comprises a template, a primer, a deoxyribonucleotide triphosphate (dNTP) and a polymerase;

the at least one conducting wire is applied with an electric current or a voltage to heat the heating coil such that buoyancy in an inside of the liquid is generated, thereby driving the analyzer to move to a top of the inside of the liquid and subsequently to move to a periphery of the inside of the liquid so as to form a thermal cycle; and

the template is amplified by recycling the thermal cycle.

10. The PCR droplet device as claimed in claim 9, wherein the first containing area is formed by a first partition surrounding a periphery of the heating coil.

11. The PCR droplet device as claimed in claim 10, wherein a distance between the first partition and the heating coil is in a range of 0.5 to 2.5 mm.

12. The PCR droplet device as claimed in claim 11, wherein the second containing area is formed by a second partition surrounding a periphery of the first partition.

13. The PCR droplet device as claimed in claim 12, wherein a distance between the second partition and the heating coil is in a range of 1 to 3 mm.

14. The PCR droplet device as claimed in claim 9, wherein the first containing area is coated with a hydrophilic material.

15. The PCR droplet device as claimed in claim 14, wherein a functional group of the hydrophilic material comprises a hydrogen group, a carbonyl group, a carboxyl group, hydroxyl group, a sulfonic acid group, or an amino group.

16. The PCR droplet device as claimed in claim 15, wherein the second containing area is coated with a hydrophobic material.

17. The PCR droplet device as claimed in claim 16, wherein the hydrophobic material comprises epoxide, cyclic acetal, or styrene.

18. The PCR droplet device as claimed in claim 9, wherein the hydrophobic solution comprises mineral oil.

19. The PCR droplet device as claimed in claim 9, wherein a material of the substrate comprises silica, glass, nylon, polymer, or ceramic.

20. The PCR droplet device as claimed in claim 9, wherein a metal electrical conductivity of the heating coil is higher than a metal electrical conductivity of the at least one conducting wire.

21. The PCR droplet device as claimed in claim 20, wherein a material of the heating coil comprises silver, copper, gold, platinum, aluminum, iron, stannum, lead or a combination thereof.

22. The PCR droplet device as claimed in claim 21, wherein a material of the at least one conducting wire comprises silver, copper, gold, platinum, aluminum, iron, stannum, lead or a combination thereof.

23. The PCR droplet device as claimed in claim 9, wherein the template comprises deoxyribonucleic acid (DNA) or ribonucleic acid (RNA).

24. The PCR droplet device as claimed in claim 9, wherein a shape of the heating coil comprises a circle, an angle, or an irregular shape.

25. The PCR droplet device as claimed in claim 9, wherein the liquid further comprises glycerol.

26. A polymerase chain reaction (PCR) droplet device array comprising a plurality of the PCR droplet device as claim 9, and the plurality of the PCR droplet devices disposed in an array mode.