THERMOCYCLER AND SAMPLE VESSEL FOR RAPID AMPLIFICATION OF DNA

• US 20110039305A1
• Filed: 02/19/2009
• Published: 02/17/2011
• Est. Priority Date: 02/20/2008
• Status: Active Grant

• Current Assignee: Streck
• Original Assignee: Streck
• Inventors: Whitney, Scott E., Termaat, Joel R., Viljoen, Hendrik J.
• Granted Patent: US 9,034,635 B2
• Time in Patent Office: Days
• US Class Current: 435/91.2
• CPC Class Codes: B01L 2300/043 Hinged closures
• CPC Class Codes: B01L 2300/0627 Sensor or part of a sensor is integrated
• CPC Class Codes: B01L 2300/0838 Capillaries
• CPC Class Codes: B01L 2300/18 Means for temperature control
• CPC Class Codes: B01L 2300/1822 using Peltier elements
• CPC Class Codes: B01L 2300/1844 using fans
• CPC Class Codes: B01L 3/505 flexible containers not provided for above
• 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

A thermocycler apparatus and method for rapidly performing the PCR process employs at least two thermoelectric modules which are in substantial spatial opposition with an interior space present between opposing modules. One or multiple sample vessels are placed in between the modules such that the vessels are subjected to temperature cycling by the modules. The sample vessels have a minimal internal dimension that is substantially perpendicular to the modules that facilitates rapid temperature cycling. In embodiments of the invention the sample vessels may be deformable between: a) a shape having a wide mouth to facilitate filling and removing of sample fluids from the vessel, and b) a shape which is thinner for conforming to the sample cavity or interior space between the thermoelectric modules of the thermocycler for more rapid heat transfer.

Claims

1. A thermocycler for subjecting one or a plurality of samples to rapid thermal cycling comprising:

at least one pair of thermoelectric modules each having an interior module face for heating and cooling one or a plurality of sample vessels each containing a sample;

wherein the thermoelectric modules of each pair are positioned such that;

the module faces of the thermoelectric module pair are in substantial opposition to each other with an interior sample holder between the opposing module faces for receiving said one or a plurality of sample vessels; and

a controller electrically connected to each pair of thermoelectric modules for regulating the temperature so that any sample vessels placed within the sample holder experience uniform temperature cyclings;

wherein the sample holder includes one or more openings for receiving the sample vessel, each opening having a shape so that the sample vessel is deformed upon entry into the sample holder openings.

27. A thermocycler as claimed in claim 1, wherein the thermoelectric modules of each pair are positioned such that the module faces of each thermoelectric module pair are in substantial opposition such that the semiconductor elements in the opposing modules are separated by a distance of about 0.5 mm to about 10.0 mm.

28. A thermocycler as claimed in claim 27, having a sample vessel wherein the distance between inner opposing surfaces of said sample vessel is from about 0.4 mm to about 2.5 mm in a direction substantially perpendicular to the opposing module faces.

38. The thermocycler of claim 1, wherein the thermoelectric modules are the only provided sources of heating or cooling the sample vessels.

39. The thermocycler of claim 1, wherein the sample vessel includes a polypropylene material.

40. The thermocycler of claim 1, wherein the sample holder is composed of a solid material.

41. The thermocycler of claim 1, wherein the sample holder includes a silver material.

42. The thermocycler of claim 1, wherein the sample vessel is substantially deformed to fit into a substantially oval shaped opening within the sample holder so that the distance between one or more walls of the sample vessel is minimized.

43. The thermocycler of claim 1, wherein the sample holder includes a plurality of oval bores.

44. The thermocycler of claim 1, wherein the sample holder includes a sensor in which the voltage or resistance signal changes with temperature to measure the temperature within the sample holder.

45. The thermocycler of claim 1, which is capable of amplifying an about 163 base pair sample located within the deformable sample vessel when subjected to 30 amplification cycles in about 300 seconds as analyzed by gel electrophoresis.

46. The thermocycler of claim 1, which is capable of amplifying an about 402 base pair sample located within the deformable sample vessel when subjected to 30 amplification cycles in about 517 seconds as analyzed by gel electrophoresis.

47. The thermocycler of claim 1, which is capable of processing a sample of from about 25 μ

l to about 250 μ

l is amplified by the thermocycler in cycle times of about 2 seconds to about 20 seconds and provides accurate gel electrophoresis results for the product amplified.

48. The thermocycler of claim 1, which is capable of processing a sample of about 100 μ

l through a PCR cycle spanning 94°

C. to 60°

C. in about 9 seconds.

49. The thermocycler of claim 1, wherein the sample vessel can hold contents of about 10 μ

l to about 250 μ

l in volume, the temperature of a sample can be varied between a low temperature range of about 55°

C. to about 72°

C. and a high temperature range of about 85°

C. to about 98°

C. and back to the low temperature range in a time frame of from about 2 seconds to about 20 seconds per cycle.

50. The thermocycler of claim 1, wherein the sample vessel is substantially deformable between a first sample filling shape prior to insertion into the sample holder and a second rapid thermocycling shape after insertion into the sample holder.

51. The thermocycler of claim 1, wherein the thermoelectric modules are hinged together at one end for insertion and removal of sample vessels from the sample holder when the hinge is opened, and thermocycling when the hinge is closed.

52. The thermocycler of claim 1, including at least two pairs of thermoelectric modules, wherein the controller controls the pairs of thermoelectric modules so that the modules run independent temperature protocols simultaneously.

53. The thermocycler of claim 1, wherein the thermoelectric modules are positioned such that the module faces of each thermoelectric module pair are in substantial opposition such that semiconductor elements in the opposing modules are separated by a distance of about 0.5 mm to about 10.0 mm.

2-26. -26. (canceled)

29. A method for performing a polymerase chain reaction comprising:

a) adding components for a polymerase chain reaction into a sample vessel, the vessel being in a sample filling configuration;

b) inserting the sample vessel into a thermocycler having a sample holder adapted to receive the sample vessel between a pair of thermoelectric modules so that at least a portion of the sample vessel deforms into a rapid cycling configuration; and

c) cycling the temperature of the vessel contents by direct or indirect contact of the vessel with the thermoelectric modules.

31. A method as claimed in claim 29 wherein the temperature of the contents of the sample vessel is cycled between a low temperature range of about 55°

C. to about 72°

C. and a high temperature range of about 85°

C. to about 98°

C. and back to the low temperature range in a time frame of from about 2 seconds to about 20 seconds per cycle.

32. A method as claimed in claim 29 wherein the temperature of the contents of the sample vessel is cycled to synthesize copies of DNA of from about 50 to about 1.000 nucleic acid base pairs in length by the polymerase chain reaction.

33. A method as claimed in claim 29 wherein sample volumes of from about 10 μ

L to about 250 μ

L are employed in the sample vessel.

54. The method of claim 29, wherein the thermoelectric modules are the only provided sources of heating or cooling the sample vessels.

55. The method of claim 29, wherein the temperature of the contents of the sample vessel is cycled between a low temperature range of about 55°

C. to about 72°

C. and a high temperature range of about 85°

C. to about 98°

C. and back to the low temperature range in a time frame of from about 2 seconds to about 20 seconds per cycle.

56. The method of claim 29, wherein the temperature of the contents of the sample vessel is cycled to synthesize copies of DNA of from about 50 to about 1,000 nucleic acid base pairs in length by the polymerase chain reaction.

57. The method of claim 29, wherein the sample holder is composed of a solid material.

58. The method of claim 29, wherein the sample holder includes a silver material.

30. (canceled)

34-37. -37. (canceled)

59. A thermocycler for subjecting one or a plurality of samples to rapid thermal cycling comprising:

a heat source consisting essentially of at least one pair of thermoelectric modules each having an interior module face for heating and cooling one or a plurality of sample vessels each containing a sample;

wherein the thermoelectric modules of each pair are positioned so that;

the module faces of the thermoelectric module pair are in substantial opposition to each other with a sample holder composed of a, solid material having a high thermal conductivity but low thermal mass between the opposing module faces for receiving said one or a plurality of sample vessels, and;

a controller electrically connected to each pair of thermoelectric modules for regulating the temperature so that any sample vessels placed within the sample holder experience, uniform temperature cycling.

60. The thermocycler of claim 59, wherein the sample holder includes a silver material.

61. The thermocycler of claim 59, wherein the temperature within the sample holder is cycled between a low temperature range of about 55°

C. to about 72°

C. and a high temperature range of about 85°

C. to about 98°

C. and back to the low temperature range in a time frame of from about 2 seconds to about 20 seconds per cycle.

62. The thermocycler of claim 59, which is capable of amplifying an about 163 base pair sample located within the deformable sample vessel when subjected to 30 amplification cycles in about 300 seconds as analyzed by gel electrophoresis.

63. The thermocycler of claim 59, which is capable of amplifying an about 402 base pair sample located within the deformable sample vessel when subjected to 30 amplification cycles in about 517 seconds as analyzed by gel electrophoresis.

64. The thermocycler of claim 59, which is capable of processing a sample of from about 25 μ

l to about 250 μ

l is amplified by the thermocycler in cycle times of about 2 seconds to about 20 seconds and provides accurate gel electrophoresis results for the product amplified.

65. The thermocycler of claim 59, which is capable of processing a sample of about 100 μ

l through a PCR cycle spanning 94°

C. to 60°

C. in about 9 seconds.

66. The thermocycler of claim 59, wherein the sample holder includes a silver material and the sample vessel is a glass capillary.

67. The thermocycler of claim 59, wherein the sample vessel can hold contents of about 10 μ

l to about 250 μ

l in volume, the temperature of a sample can be varied between a low temperature range of about 55°

C. to about 72°

C. and a high temperature range of about 85°

C. to about 98°

C. and back to the low temperature range in a time frame of from about 2 seconds to about 20 seconds per cycle.

68. The thermocycler of claim 59, wherein the sample vessel is resilient forming a first shape prior to insertion into the sample holder, a second shape after insertion into the sample holder, and returning to substantially the first shape after removal from the sample holder.

69. The thermocycler of claim 59, wherein the thermoelectric modules are hinged together at one end for insertion and removal of sample vessels from the sample holder when the hinge is opened, and thermocycling when the hinge is closed.

70. The thermocycler of any claim 59, including at least two pairs of thermoelectric modules, wherein the controller controls the pairs of thermoelectric modules so that the modules run independent temperature protocols simultaneously.