Device for the amplification of dna, comprising a microwave energy source

• US 20050233324A1
• Filed: 05/01/2003
• Published: 10/20/2005
• Est. Priority Date: 05/01/2002
• Status: Abandoned Application

• Current Assignee: Corbett Life Science (Qiagen NV)
• Original Assignee: Corbett Life Science (Qiagen NV)
• Inventors: Corbett, John Michael, Corbett, John Michael Jr.
• Application Number:
  US10/512,972
• Publication Number:
  US 20050233324A1
• Time in Patent Office: Days
• US Class Current: 435/6
• CPC Class Codes: B01L 2200/147 Employing temperature sensors
• CPC Class Codes: B01L 2300/0627 Sensor or part of a sensor is integrated
• CPC Class Codes: B01L 2300/0803 Disc shape
• CPC Class Codes: B01L 2300/1844 using fans
• CPC Class Codes: B01L 2300/1866 Microwaves
• 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: B01L 7/5255 by moving sample containers
• CPC Class Codes: B04B 15/02 for cooling, heating, or heat insulating
• CPC Class Codes: C12Q 1/6816 characterised by the detection means C12Q1/6804 takes precedence
• CPC Class Codes: C12Q 1/686 Polymerase chain reaction [PCR]
• CPC Class Codes: C12Q 2523/313 Irradiation, e.g. UV irradiation
• CPC Class Codes: C12Q 2563/173 staining/intercalating agent, e.g. ethidium bromide
• CPC Class Codes: G01N 2035/00405 Microwaves
• CPC Class Codes: G01N 2035/0444 for cuvettes or reaction vessels

Abstract

The invention relates to a device for the amplification of DNA in a reaction mixture, the device (1) comprising a heated chamber (2) including a rotor (3) for holding a plurality of reaction vessels for reaction mixtures, a drive means for the rotor, a microwave energy source (8) with means for controlled delivery of said energy to the reaction mixtures, and a system (14-16) for determining denaturation of double-stranded DNA. The invention also provides a method for the amplification of a nucleic acid strand. In the first step of the method, a reaction mixture is formed comprising the target nucleic acid strand, nucleotides, a primer, a thermostable nucleic acid polymerase, and, if necessary, a reagent for the detection of denaturation of double-stranded DNA. In the second step, the mixture is incubated at a temperature which allows synthesis of a nucleic acid strand complementary to the target nucleic acid strand. The third step comprises denaturing double-stranded DNA formed in the second step by microwave energisation of the reaction mixture with monitoring of the mixture to determine the denaturation end point. The reaction mixture is allowed to cool to a temperature at which primer anneals to the target nucleic acid strand in the fourth step. The second to fourth steps are repeated until a desired level of amplification is achieved.

Claims

1. A device for the amplification of DNA in a reaction mixture, the device comprising:

a heated chamber including a rotor for holding a plurality of reaction vessels for reaction mixtures;

a drive means for said rotor;

a microwave energy source with means for controlled delivery of said energy to said reaction mixtures; and

a system for determining denaturation of double-stranded DNA.

2. The device of claim 1, wherein said chamber has a lid or sealable opening for loading the device rotor.

3. The device of claim 1, wherein said heating of said chamber is by an internal heater or by supplying heated air through at least one port in a wall of said chamber.

4. The device of claim 1, which further includes temperature sensor within the chamber which is linked to an associated computer responsible for controlling the operation of the device.

5. The device of claim 1, wherein said rotor is fabricated from a plastics material.

6. The device of claim 1, wherein said rotor drive means is a direct-coupled AC motor, DC motor or stepper motor.

7. The device of claim 1, wherein said microwave energy source comprises a magnetron that is external to the device chamber with energy delivered to reaction vessels via a wave guide.

8. The device of claim 1, wherein said system for determining denaturation of double-stranded DNA is a fluorescence detection system or an infrared measurement system.

9. The device of claim 1, which further includes a mechanism for cooling the device chamber.

11. The method of claim 10, wherein said method is carried out in a device according to claim 1.

10. A method for the amplification of a nucleic acid strand, the method comprising the steps of:

i) forming a reaction mixture comprising said target nucleic acid strand, nucleotides, a primer, a thermostable nucleic acid polymerase, and, if necessary, a reagent for the detection of denaturation of double-stranded DNA;

ii) incubating said mixture at a temperature which allows synthesis of a nucleic acid strand complementary to said target nucleic acid strand;

iii) denaturing double-stranded DNA formed in step (ii) by microwave energisation of said reaction mixture with monitoring of said mixture to determine the denaturation end point;

iv) allowing said reaction mixture to cool to a temperature at which primer anneals to said target nucleic acid strand; and

v) repeating steps (ii) to (iv) until a desired level of amplification is achieved.

12. The method of claim 10, wherein said reagent for the detection of the denaturation of double stranded DNA is a fluorophore.