ASSAY AND OTHER REACTIONS INVOLVING DROPLETS
First Claim
1. A method of polynucleotide amplification comprising:
- a. generating a library of droplets comprising a plurality of polynucleotide primers, wherein the library comprises more than 105 polynucleotide primers;
b. generating a set of droplets, wherein the droplets comprise at least one polynucleotide to be amplified;
c. merging a first droplet from the library of droplets with a second droplet from the set of droplets to form a coalesced droplet; and
d. performing an amplification reaction on at least one polynucleotide in the coalesced droplet to form at least one amplicon.
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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
21 Claims
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1. A method of polynucleotide amplification comprising:
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a. generating a library of droplets comprising a plurality of polynucleotide primers, wherein the library comprises more than 105 polynucleotide primers; b. generating a set of droplets, wherein the droplets comprise at least one polynucleotide to be amplified; c. merging a first droplet from the library of droplets with a second droplet from the set of droplets to form a coalesced droplet; and d. performing an amplification reaction on at least one polynucleotide in the coalesced droplet to form at least one amplicon. - View Dependent Claims (2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21)
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Specification