Assays and other reactions involving droplets
First Claim
1. A method for nucleic acid processing, comprising:
- (a) directing (i) a first phase comprising reagents, and a plurality of cells, and (ii) a second phase comprising an oil to an intersection of a plurality of channels, wherein said first phase is immiscible with said second phase;
(b) at said intersection, generating a plurality of droplets comprising said plurality of cells and said reagents, wherein an individual droplet of said plurality of droplets comprises a single cell of said plurality of cells, said single cell comprising a first polynucleotide comprising a nucleic acid sequence;
(c) subjecting said first polynucleotide to release from said single cell; and
(d) using said reagents, generating from said first polynucleotide a second polynucleotide comprising a sequence derived from said nucleic acid sequence.
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Accused Products
Abstract
The present invention generally relates to droplets and/or emulsions, such as multiple emulsions. In some cases, the droplets and/or emulsions may be used in assays, and in certain embodiments, the droplet or emulsion may be hardened to form a gel. In some aspects, a heterogeneous assay can be performed using a gel. For example, a droplet may be hardened to form a gel, where the droplet contains a cell, DNA, or other suitable species. The gel may be exposed to a reactant, and the reactant may interact with the gel and/or with the cell, DNA, etc., in some fashion. For example, the reactant may diffuse through the gel, or the hardened particle may liquefy to form a liquid state, allowing the reactant to interact with the cell. As a specific example, DNA contained within a gel particle may be subjected to PCR (polymerase chain reaction) amplification, e.g., by using PCR primers able to bind to the gel as it forms. As the DNA is amplified using PCR, some of the DNA will be bound to the gel via the PCR primer. After the PCR reaction, unbound DNA may be removed from the gel, e.g., via diffusion or washing. Thus, a gel particle having bound DNA may be formed in one embodiment of the invention.
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Citations
33 Claims
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1. A method for nucleic acid processing, comprising:
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(a) directing (i) a first phase comprising reagents, and a plurality of cells, and (ii) a second phase comprising an oil to an intersection of a plurality of channels, wherein said first phase is immiscible with said second phase; (b) at said intersection, generating a plurality of droplets comprising said plurality of cells and said reagents, wherein an individual droplet of said plurality of droplets comprises a single cell of said plurality of cells, said single cell comprising a first polynucleotide comprising a nucleic acid sequence; (c) subjecting said first polynucleotide to release from said single cell; and (d) using said reagents, generating from said first polynucleotide a second polynucleotide comprising a sequence derived from said nucleic acid sequence. - View Dependent Claims (2, 3, 4, 5, 6, 18, 19, 20, 21, 23, 25, 27, 30, 31, 32)
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7. A method for nucleic acid processing, comprising:
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(a) generating a plurality of droplets each comprising a particle comprising a primer sequence, a single cell comprising a first polynucleotide having a nucleic acid sequence, and reagents; (b) subjecting said first polynucleotide to release from said single cell; and (c) using said primer sequence and said reagents, generating from said first polynucleotide a second polynucleotide comprising a sequence derived from said nucleic acid sequence. - View Dependent Claims (8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 22, 24, 26, 28, 29, 33)
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Specification