Nucleic acid amplification reaction apparatus and method

• US 5,779,977 A
• Filed: 03/26/1997
• Issued: 07/14/1998
• Est. Priority Date: 07/28/1993
• Status: Expired due to Term

• Current Assignee: Applied Biosystems LLC (Thermo Fisher Scientific Incorporated)
• Original Assignee: The Perkin-Elmer Corporation (Thermo Fisher Scientific Incorporated)
• Inventors: Woudenberg, Timothy M., Haff, Lawrence A., Bloch, Will, Picozza, Enrico
• Primary Examiner(s): Marschel, Ardin H.
• Application Number:
  US08/827,353
• Time in Patent Office: 475 Days
• Field of Search: 422/50, 422/62, 422/68.1, 422/82.05, 422/99, 422/129, 422/131, 422/132, 422/134, 422/138, 422/149, 422/188, 422/198, 422/202, 422/203, 435/91.1, 435/91.2, 435/91.5, 435/283.1, 435/286.1, 435/286.5, 435/289.1, 435/293.1, 435/303.1, 436/501, 935/85, 935/88
• US Class Current: 422/68.1
• CPC Class Codes: B01J 19/0046 Sequential or parallel reactions, e.g. for the synthesis of polypeptides or polynucleotides; Apparatus and devices for combinatorial chemistry or for making molecular arrays synthesis methods per se C40B50/00
• CPC Class Codes: B01J 2219/00495 Means for heating or cooling the reaction vessels
• CPC Class Codes: B01J 2219/00722 Nucleotides
• CPC Class Codes: B01L 2300/1883 using thermal insulation
• 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 7/525 with physical movement of samples between temperature zones
• CPC Class Codes: B01L 9/065 specially adapted for capillary tubes
• CPC Class Codes: C12Q 1/686 Polymerase chain reaction [PCR]
• CPC Class Codes: C40B 40/06 Libraries containing nucleotides or polynucleotides, or derivatives thereof
• CPC Class Codes: C40B 60/14 for creating libraries
• CPC Class Codes: G01N 2035/00237 Handling microquantities of analyte, e.g. microvalves, capillary networks
• CPC Class Codes: Y10T 436/116664 with automated titrator

Abstract

Apparatus and method for performing a nucleic acid amplification reaction and preferably a polymerase chain reaction (PCR) in a reaction mixture in at least one capillary tube. Several different embodiments are disclosed. One embodiment cycles a sample through a capillary tube loop passing through two thermostatted fluid baths. Another embodiment has the capillary tube routed alternatingly between two heat exchangers to that the sample makes only one pass through the tube. Other embodiments maintain the heat exchangers stationary and translate the samples between them. Still further embodiments maintain the samples stationary and either automatically translate or rotate the heat exchangers past the samples contained within the capillary tubes to perform the thermal cycles necessary for the amplification reaction.

Claims

1. An apparatus for performing a nucleic acid amplification reaction in a reaction mixture, comprising:

a capillary tube for containing the reaction mixture;

a chamber surrounding the capillary tube including at least one input port for receiving fluid of different temperatures and at least one output port through which the fluid can be removed from the chamber;

a first thermostatted bath coupled to the chamber through the input and output ports, the bath containing a temperature-stabilized body of fluid at a temperature suitable for causing nucleic acid denaturation to occur in the reaction mixture;

a second thermostatted bath coupled to the input and output ports of the chamber containing a temperature-stabilized body of fluid at a temperature suitable for causing nucleic acid annealing and extension to occur in the reaction mixture; and

pumping means for alternately moving fluid from the first and second baths into and out of the chamber so as to alternatingly subject the reaction mixture to the denaturation and annealing temperatures.

2. The apparatus of claim 1 wherein the pumping means comprises a pump in each of the first and second baths, each of which is coupled to separate input and output ports of the chamber, and each of which is controlled by a computer directed controller for selectively pumping fluids into and out of the chamber.

3. The apparatus of claim 2 wherein the pumping means is a single, shared, bidirectional pump coupled to a port of the chamber and coupled to a fluid multiplexer, the multiplexer having one bi-directional port coupled to the first bath and a second bi-directional port coupled to the second bath for selectively coupling a selectable one of the baths to the pump, the pump and multiplexer being under the control of the computer-directed controller for controlling the pump and the fluid multiplexer so as to cyclically and alternately selectively pumping fluid from the first and second baths into the chamber to implement a nucleic acid amplification reaction protocol defined by stored data in the computer directed controller.

4. An apparatus for performing a chemical chain reaction in a capillary tube, comprising:

an elongated chamber having an input port and an output port and having a wire mesh across at least a cross sectional portion of the chamber;

at least first and second fluid bypass pipes providing fluid flow paths around the chamber;

a first thermostatted bath for maintaining the temperature of a body of fluid therein at a temperature suitable for causing denaturation to occur in a reaction mixture;

a second thermostatted bath for maintaining the temperature of a body of fluid therein at a temperature suitable for causing annealing and extension reactions to occur in the reaction mixture;

pumping means connected between the chamber and each of the baths for pumping fluid from each of the first and second baths to the chamber;

valve means for selectively coupling a stream of fluid from one of the baths through the chamber while coupling another stream of fluid from the second bath through one of the bypass pipes, and then for coupling the stream pumped by the pumping means from the second bath through the chamber and coupling the stream pumped by the pumping means from the first bath through another one of the bypass pipes.