Mesoscale polynucleotide amplification device and method

Abstract

Disclosed are devices for amplifying a preselected polynucleotide in a sample by conducting a polynucleotide polymerization reaction. The devices comprise a substrate microfabricated to define a sample inlet port and a mesoscale flow system, which extends from the inlet port. The mesoscale flow system includes a polynucleotide polymerization reaction chamber in fluid communication with the inlet port which is provided with reagents required for polymerization and amplification of a preselected polynucleotide. In one embodiment the devices may be utilized to implement a polymerase chain reaction (PCR) in the reaction chamber (PCR chamber). The PCR chamber is provided with the sample polynucleotide, polymerase, nucleoside triphosphates, primers and other reagents required for the polymerase chain reaction, and the device is provided with means for thermally controlling the temperature of the contents of the reaction chamber at a temperature controlled to dehybridize double stranded polynucleotide, to anneal the primers, and to polymerize and amplify the polynucleotide.

1 . an analytical system for PCR analysis comprising: 
 a device comprising a mesoscale flow system and an inlet port communicating with the flow system;    an appliance comprising a nesting site that releasably holds the device, a flow line mating with the inlet port of the device, a pump connected to the flow line that when actuated forces fluid through the flow system of the device, and a sensor that detects properties of fluid within the flow system; and    means for thermally cycling fluid within the flow system.    
     
     
         2 . The system of  claim 1 , wherein the device comprises a solid substrate.  
     
     
         3 . The system of  claim 2 , wherein the solid substrate is approximately 0.2 to 2.0 centimeters square.  
     
     
         4 . The system of  claim 2 , wherein the device further comprises a cover.  
     
     
         5 . The system of  claim 4 , wherein the cover is a transparent cover.  
     
     
         6 . The system of  claim 2 , wherein the solid substrate comprises silicon.  
     
     
         7 . The system of  claim 2 , wherein the solid substrate comprises a polymer.  
     
     
         8 . The system of  claim 1 , wherein the mesoscale flow system comprises a flow channel extending from the inlet port and in fluid communication with a reaction chamber.  
     
     
         9 . The system of  claim 8 , wherein the reaction chamber has a larger cross-sectional dimension than the flow channel.  
     
     
         10 . The system of  claim 8 , wherein the reaction chamber has a cross-sectional width of 1-5 mm.  
     
     
         11 . The system of  claim 9 , wherein the reaction chamber has a cross-sectional depth of 2-50 μm.  
     
     
         12 . The system of  claim 1 , wherein the sensor is capable of detecting changes in optical properties of fluid within the flow system.  
     
     
         13 . The system of  claim 12 , wherein the sensor is a spectrophotometer.  
     
     
         14 . The system of  claim 12 , wherein the sensor comprises a video camera.  
     
     
         15 . The system of  claim 1 , wherein the sensor is a pressure detector.  
     
     
         16 . The system of  claim 1 , wherein the sensor is an electrical conductivity sensor.  
     
     
         17 . The system of  claim 1 , wherein the appliance comprises a magnetic source.  
     
     
         18 . The system of  claim 1 , wherein the appliance comprises electrical contacts in the nesting site, the device comprises electrical contacts, and the electrical contacts in the appliance mate with electrical contacts on the device when the device is placed in the nesting site.  
     
     
         19 . The system of  claim 18 , wherein the appliance is configured to transmit electrical energy to the device.  
     
     
         20 . The system of  claim 18 , wherein the appliance is configured to receive electrical signals from the device.  
     
     
         21 . The system of  claim 1 , wherein the nesting site comprises a heating element.  
     
     
         22 . The system of  claim 1 , wherein the nesting site comprises a cooling element.  
     
     
         23 . The system of  claim 1 , wherein the device comprises multiple inlet ports.  
     
     
         24 . The system of  claim 8 , wherein the means for thermally cycling fluid comprises a heating/cooling element within the reaction chamber.  
     
     
         25 . The system of  claim 19 , wherein the device comprises a solid substrate, and wherein the means for thermally cycling fluid comprises an electrical heating element integrated into the substrate.  
     
     
         26 . The system of  claim 1 , wherein the sensor is a thermocouple.