ASSAYS AND OTHER REACTIONS INVOLVING DROPLETS
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.
1 - 20 . (canceled)
21 . A method, comprising:
providing a plurality of droplets comprising cells, wherein the cells comprise polynucleotides;
releasing the polynucleotides from the cells; and
generating additional polynucleotides from the released polynucleotides within the plurality of droplets.
22 . The method of claim 21 , further comprising subjecting said plurality of droplets to conditions sufficient to generate gels in said plurality of droplets, which gels comprise polynucleotides.
23 . The method of claim 21 , wherein generating additional polynucleotides from the released polynucleotides comprises performing polymerase chain reaction (PCR).
24 . The method of claim 21 , wherein generating additional polynucleotides from the released polynucleotides comprises performing a reverse transcription reaction.
25 . The method of claim 21 , further comprising sequencing the additional polynucleotides.
26 . A method, comprising:
providing a plurality of droplets comprising a plurality of cells, wherein a droplet of the plurality of droplets comprises a cell of the plurality of cells, wherein the cells comprise polynucleotides;
releasing the polynucleotides from the cells; and
performing a polymerase chain reaction (PCR) to generate amplified polynucleotides within the droplets.
27 . The method of claim 26 , further comprising subjecting said plurality of droplets to conditions sufficient to generate gels in said plurality of droplets, which gels comprise polynucleotides.
28 . The method of claim 26 , further comprising sequencing the amplified polynucleotides.
29 . A method for nucleic acid processing comprising:
providing a plurality of droplets comprising a plurality of cells and reagents, wherein an individual droplet of said plurality of droplets comprises a single cell of said plurality of cells, the single cell comprising a first polynucleotide comprising a nucleic acid sequence,
releasing the first polynucleotide from the single cell; and
generating a second polynucleotide from the first polynucleotide comprising a sequence derived from the nucleic acid sequence.
30 . The method of claim 29 , further comprising sorting the plurality of droplets to enrich for the individual droplet.
31 . The method of claim 29 , wherein the single cell is in a gel capsule.
32 . The method of claim 29 , further comprising subjecting the plurality of droplets to conditions sufficient to generate gels in the plurality of droplets, which gels comprise polynucleotides, including the first polynucleotide.
33 . The method of claim 29 , wherein amplifying the first polynucleotide to generate a second polynucleotide comprises performing polymerase chain reaction (PCR) on the first polynucleotide.
34 . The method of claim 29 , wherein generating additional polynucleotides from the released polynucleotides comprises performing a reverse transcription reaction.
35 . The method of claim 29 , further comprising sequencing the second polynucleotide or a derivative thereof.
35 . A method for nucleic acid processing comprising:
providing a plurality of droplets comprising a plurality of cells and reagents, wherein an individual droplet of the plurality of droplets comprises a single cell of the plurality of cells, the single cell comprising a first polynucleotide comprising a nucleic acid sequence;
releasing the first polynucleotide from the single cell; and
performing a polymerase chain reaction (PCR) to amplify the first polynucleotide to generate a second polynucleotide comprising a sequence derived from the nucleic acid sequence.
36 . The method of claim 35 , further comprising sorting the plurality of droplets to enrich for the individual droplet.
37 . The method of claim 35 , wherein the single cell is in a gel capsule.
38 . The method of claim 35 , further comprising subjecting the plurality of droplets to conditions sufficient to generate gels in the plurality of droplets, which gels comprise polynucleotides, including the first polynucleotide.
39 . The method of claim 35 , further comprising sequencing the second polynucleotide or a derivative thereof.