Microfluidic device for sequencing by hybridization
First Claim
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1. A microfluidic system, comprising:
- a set of oligonucleotides dried on one or more reagent storage substrates, the set comprising at least 70% of all possible n-mers for a length n, wherein n is at least 4;
a sample nucleic acid; and
, a material transport system configured to flow members of the set of oligonucleotides into contact with the nucleic acid sample, wherein the material transport system accesses the one or more reagent storage substrate to flow members of the set of oligonucleotides into contact with the nucleic acid sample.
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Abstract
Integrated systems, apparatus, software, and methods for performing biochemical analysis, including DNA sequencing, genomic screening, purification of nucleic acids and other biological components and drug screening are provided. Microfluidic devices, systems and methods for using these devices and systems for performing a wide variety of fluid operations are provided. The devices and systems of are used in performing fluid operations which require a large number of iterative, successive or parallel fluid manipulations, in a microscale, or sealed and readily automated format.
245 Citations
61 Claims
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1. A microfluidic system, comprising:
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a set of oligonucleotides dried on one or more reagent storage substrates, the set comprising at least 70% of all possible n-mers for a length n, wherein n is at least 4;
a sample nucleic acid; and
,a material transport system configured to flow members of the set of oligonucleotides into contact with the nucleic acid sample, wherein the material transport system accesses the one or more reagent storage substrate to flow members of the set of oligonucleotides into contact with the nucleic acid sample. - View Dependent Claims (2, 3, 4, 5, 6, 7, 8, 9, 10, 12, 13, 14, 15, 16, 17, 18, 29, 30, 31, 32, 33, 34, 35, 36)
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11. A microfluidic system, comprising:
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a set of oligonucleotides, the set comprising at least 70% of all possible n-mers for a length n wherein n is at least 4;
a sample nucleic acid;
a material transport system configured to flow members of the set of oligonucleotides into contact with the sample nucleic acid; and
a molecular beacon which hybridizes to a SNP in the sample nucleic acid.
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19. A microfluidic system, comprising:
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a set of oligonucleotides, the set comprising at least 70% of all possible n-mers for a length n, wherein n is at least 4;
a sample nucleic acid;
a material transport system configured to flow members of the set of oligonucleotides into contact with the sample nucleic acid; and
a microwell plate comprising the set of oligonucleotides and a substrate comprising at least one microchannel in or one the substrate, the material transport system comprising a capillary channel that extends outward from the substrate, wherein, during operation of the system, the capillary channel is contacted to one or more well of the microwell plate, thereby permitting the material transport system to draw one or more oligonucleotide from the microwell plate into the microchannel.
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20. A microfluidic system, comprising:
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a set of oligonucleotides, the set comprising at least 70% of all possible n-mers for a length n, wherein n is at least 4;
a sample nucleic acid;
a material transport system configured to flow members of the set of oligonucleotides into contact with the sample nucleic acid; and
a reagent substrate comprising the set of oligonucleotides in dried form on the reagent substrate and a microfluidic substrate comprising at least one microchannel in or on the microfluidic substrate, the material transport system comprising a capillary channel that extends outward from the substrate, wherein, during operation of the system, the capillary channel expels fluid from the tip of the capillary channel onto one or more oligonucleotide on the reagent substrate, thereby solubilizing the oligonucleotide and draws the resulting solubilized oligonucleotide into the capillary channel, thereby permitting the material transport system to draw one or more oligonucleotide from the reagent substrate into the microchannel.
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21. A microfluidic system, comprising:
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a set of oligonucleotides, the set comprising at least 70% of all possible n-mers for a length n, wherein n is at least 4;
a sample nucleic acid;
a material transport system configured to flow members of the set of oligonucleotides into contact with the sample nucleic acid; and
a polymerase.- View Dependent Claims (22, 23)
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24. A microfluidic system, comprising:
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a set of oligonucleotides, the set comprising at least 70% of all possible n-mers for a length n, wherein n is at least 4;
a sample nucleic acid;
a material transport system configured to flow members of the set of oligonucleotides into contact with the sample nucleic acid; and
a ligase.
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25. A microfluidic system, comprising:
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a set of oligonucleotides, the set comprising at least 70% of all possible n-mers for a length n, wherein n is at least 4;
a sample nucleic acid;
a material transport system configured to flow members of the set of oligonucleotides into contact with the sample nucleic acid; and
a ligase, which ligase ligates one or more members of the set of oligonucleotides, which oligonucleotides are ligated while hybridized to the sample nucleic acid.- View Dependent Claims (26)
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27. A microfluidic system, comprising:
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a set of oligonucleotides, the set comprising at least 70% of all possible n-mers for a length n, wherein n is at least 4;
a sample nucleic acid;
a material transport system configured to flow members of the set of oligonucleotides into contact with the sample nucleic acid; and
a temperature control element, which element is configured to direct the system to thermocycle the oligonucleotides or the sample nucleic acid, or both.- View Dependent Claims (28)
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37. A system, comprising:
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a set of oligonucleotides, the set comprising at least 70% of all possible n-mers for a length n, wherein n is at least 4;
a sample nucleic acid;
a material transport system configured to flow members of the set of oligonucleotides into contact with the sample nucleic acid; and
one or more microchannels coupled to the material transport system and one or more DNA size-separation matrices in the one or more microchannels.
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38. A method of detecting a nucleic acid in a microfluidic system, the method comprising:
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providing a set of oligonucleotides dried on one or more reagent storage substrates, the set comprising at least 70% of all possible n-mers for a length n, wherein n is at least 4;
providing a sample nucleic acid;
flowing at least one oligonucleotide member of the set into contact with the sample nucleic acid;
hybridizing the at least one oligonucleotide member and the sample nucleic acid; and
,detecting hybridization of the oligonucleotide member to the sample nucleic acid, thereby determining a sequence of nucleotides of the sample nucleic acid. - View Dependent Claims (39, 40, 41, 42, 43, 44, 45, 48, 49, 50, 51, 52, 53, 54, 59, 60, 61)
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46. A method of detecting a nucleic acid in a microfluidic system, the method comprising:
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providing a set of oligonucleotides, the set comprising at least 70% of all possible n-mers for a length n, wherein n is at least 4;
performing one or more amplification reaction to produce a sample nucleic acid;
flowing at least one oligonucleotide member of the set into contact with the sample nucleic acid;
hybridizing the at least one oligonucleotide member and the sample nucleic acid; and
detecting hybridization of the oligonucleotide member to the sample nucleic acid, thereby determining a sequence of nucleotides of the sample nucleic acid, wherein the one or more amplification reaction comprises a ligase-mediated amplification reaction. - View Dependent Claims (47)
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55. A method of detecting a nucleic acid in a microfluidic system, the method comprising:
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providing a set of oligonucleotides, the set comprising at least 70% of all possible n-mers for a length n, wherein n is at least 4;
providing a sample nucleic acid;
flowing at least one oligonucleotide member of the set into contact with the sample nucleic acid;
hybridizing the at least one oligonucleotide member and the sample nucleic acid; and
detecting hybridization of the oligonucleotide member to the sample nucleic acid, thereby determining a sequence of nucleotides of the sample nucleic acid, wherein detecting hybridization comprises performing one or more extension reaction by extending the hybridized oligonucleotide member and detecting the resulting extension product. - View Dependent Claims (56, 57)
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58. A method of detecting a nucleic acid in a microfluidic system, the method comprising:
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providing a set of oligonucleotides, the set comprising at least 70% of all possible n-mers for a length n, wherein n is at least 4;
providing a sample nucleic acid;
flowing at least one oligonucleotide member of the set into contact with the sample nucleic acid;
hybridizing the at least one oligonucleotide member and the sample nucleic acid; and
detecting hybridization of the oligonucleotide member to the sample nucleic acid, thereby determining a sequence of nucleotides of the sample nucleic acid, wherein detecting hybridization comprises amplifying the sample nucleic acid by extending the oligonucleotide member with a polymerase in one or more PCR or LCR reaction.
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Specification