Mesoscale polynucleotide amplification device and method
First Claim
1. A device for amplifying a preselected polynucleotide in a sample by conducting a polynucleotide amplification reaction, the device comprising:
- a solid substrate which is microfabricated to define;
a sample inlet port;
a mesoscale flow system comprising;
a sample flow channel extending from said inlet port; and
a polynucleotide amplification reaction chamber, in fluid communication with said flow channel;
said sample flow channel and said reaction chamber having at least one cross-sectional dimension of width or depth which is about 0.1 to 500 μ
m, said chamber and said channel being of dissimilar dimension; and
a fluid exit port in fluid communication with said flow system; and
means for thermally cycling the contents of said chamber whereby in each cycle the temperature is controlled to dehybridize double stranded polynucleotide and to permit the in vitro amplification of a preselected polynucleotide.
0 Assignments
0 Petitions
Accused Products
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.
-
Citations
27 Claims
-
1. A device for amplifying a preselected polynucleotide in a sample by conducting a polynucleotide amplification reaction, the device comprising:
a solid substrate which is microfabricated to define; a sample inlet port; a mesoscale flow system comprising; a sample flow channel extending from said inlet port; and a polynucleotide amplification reaction chamber, in fluid communication with said flow channel; said sample flow channel and said reaction chamber having at least one cross-sectional dimension of width or depth which is about 0.1 to 500 μ
m, said chamber and said channel being of dissimilar dimension; anda fluid exit port in fluid communication with said flow system; and means for thermally cycling the contents of said chamber whereby in each cycle the temperature is controlled to dehybridize double stranded polynucleotide and to permit the in vitro amplification of a preselected polynucleotide. - View Dependent Claims (2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13)
-
14. A device for amplifying a preselected polynucleotide in a sample, the device comprising:
-
a solid substrate microfabricated to define; a sample inlet port; a mesoscale flow system comprising; a sample flow channel extending from said inlet port; and a polynucleotide amplification reaction chamber, in fluid communication with said flow channel, containing a preselected polynucleotide and polynucleotide amplification reagents; said sample flow channel and said reaction chamber having at least one cross-sectional dimension of width or depth which is about 0.1 to 500 μ
m, said chamber and said channel being of dissimilar dimension; anda fluid exit port in fluid communication with said flow system; and means for thermally cycling the contents of said chamber whereby, in each cycle, temperature is controlled to dehybridize double stranded polynucleotide, and to permit synthesis of polynucleotide, thereby to amplify said preselected polynucleotide. - View Dependent Claims (15, 16, 17, 18, 19, 20)
-
-
21. A method for amplifying a preselected polynucleotide in a sample by conducting a polynucleotide amplification reaction, the method comprising:
-
(i) providing a device comprising; a solid substrate microfabricated to define; a sample inlet port; a mesoscale flow system comprising; a sample flow channel extending from said inlet port; and a polynucleotide amplification reaction chamber in fluid communication with said flow channel; said sample flow channel and said reaction chamber having at least one cross-sectional dimension of width or depth which is about 0.1 to 500 μ
m, said chamber and said channel being of dissimilar dimension; anda fluid exit port in fluid communication with said flow system; and means for thermally regulating the contents of said chamber at a temperature controlled to permit amplification of said preselected polynucleotide; (ii) delivering, through said inlet port and said mesoscale flow system to said reaction chamber, a sample polynucleotide and reagents required for an in vitro polynucleotide amplification reaction; and (iii) thermally controlling the contents of said chamber to permit amplification of said polynucleotide.
-
-
22. The method of claim 31 wherein said amplification reaction is a polymerase chain reaction (PCR);
-
wherein, in step (i), said means for thermally controlling comprises means for thermally cycling the contents of said chamber; wherein step (ii) includes the step of adding to said amplification chamber;
a polymerase, nucleoside triphosphates, a first primer hybridizable with said sample polynucleotide, and a second primer hybridizable with a nucleic acid comprising a sequence complementary to said polynucleotide, and wherein said first and second primers define the termini of the polynucleotide product of the polymerization reaction; andwherein step (iii) includes the step of thermally cycling the contents of said chamber whereby, in each cycle, the temperature is controlled to dehybridize double stranded polynucleotide thereby to produce single stranded polynucleotide, to permit annealing of complementary regions of single stranded polynucleotide, and to permit synthesis and polymerization of polynucleotide between said primers. - View Dependent Claims (23, 24)
-
-
25. A device for amplifying a preselected polynucleotide in a sample by conducting a polynucleotide amplification reaction, the device comprising:
a solid substrate microfabricated to define; a sample inlet port; a mesoscale flow system comprising; a sample flow channel extending from said inlet port; and a polynucleotide amplification chamber, in fluid communication with said flow channel, said chamber containing reagents for amplifying a preselected polynucleotide in vitro; said sample flow channel and said reaction chamber having at least one cross-sectional dimension of width or depth which is about 0.1 to 500 μ
m, said chamber and said channel being of dissimilar dimension; anda fluid exit port in fluid communication with said flow system. - View Dependent Claims (26, 27)
Specification