Abstract: This disclosure provides methods and systems for single-cell analysis, including single-cell transcriptome analysis, of target cells without microfluidic devices. The disclosed methods involve the use of template particles to template the formation of monodisperse droplets to generally capture a single target cell from a population of cells in an encapsulation, derive a plurality of distinct mRNA molecules from the single target cell, and quantify the distinct mRNA molecules to generate an expression profile.

Abstract: Emulsion compositions are provided herein. Also provided herein are kits containing one or more emulsion compositions or components for making such emulsion compositions. Also provided herein are methods of using such emulsion compositions, such as for amplification of target nucleic acids in emulsion droplets.

Abstract: This invention releases to systems and methods for detecting the presence, and preferably sequence, of microbial nucleic acids from a patient sample using droplets. In particular, methods involve probing patient nucleic acid samples with capture probes that include nucleotide sequences that are highly specific to microbial nucleic acids. Complementary microbial nucleic acids present in the sample binds to the capture probes, inside droplets, and are amplified into amplicons that can be readily detected. Samples positive for microbial nucleic acids may be sequenced to identify the microbe.

Abstract: This invention relates to systems and methods for making libraries of molecularly distinct polynucleotides. In particular, methods of the invention involve randomly fragmenting nucleic acids (e.g., RNA) to create fragments with cleaved ends at random cleavage locations. Preferably, methods also include reverse transcribing the fragments of RNA in the presence of molecular diversity enhancers (i.e., short random sequences), thereby creating polynucleotides with the molecular diversity enhancers copied therein. The result is a library of polynucleotides that are uniquely identifiable based on combinations of the random cleavage locations and molecular diversity enhancers.

Abstract: Emulsion compositions are provided herein. Also provided herein are kits containing one or more emulsion compositions or components for making such emulsion compositions. Also provided herein are methods of using such emulsion compositions, such as for amplification of target nucleic acids in emulsion droplets.

Abstract: The disclosure provides methods for creating long oligonucleotide reagents that include barcodes and other element for sequencing library preparation, where the oligonucleotides are created by multiple tiers of ligation of shorter oligos. The disclosed methods work to extend short oligos that are attached to particles, thereby allowing one to create particles that carry large number of long sample preparation oligonucleotides without being required to synthesize those full-length molecules with a polymerase.

Abstract: The disclosure provides methods and systems of analyzing single cells by simultaneously separating cells into monodisperse droplets and tagging each nucleic acid molecule from the cells with barcodes unique to each droplet. The methods and systems combine template particles with a plurality of single cells in a tube, generate in the tube monodispersed droplets encapsulating a single one of the template particles and a single one of the single cells, release nucleic acid molecules from the single cells and provide each nucleic acid molecule with a barcode unique to the respective droplet. The nucleic acid molecules can then be analyzed by any known method, for example by sequencing the nucleic acid molecules.

Abstract: The methods and systems described herein provide capture template particles and an improved emulsion droplet-based target capture and barcoding method thereof. The capture template particles and methods disclosed herein allow capturing targets of interest from biological samples, and barcoding of specific nucleic acids contained in the captured targets. The nucleic acids can be contained within living or nonliving structures, including particles, viruses, and cells. The nucleic acids can include, e.g., DNA or RNA.

Abstract: Microfluidic methods of altering the spacing of a stream of objects. In an exemplary method, objects of the object stream may be transported in carrier fluid along a microfluidic channel structure having an inflow region, an outflow region, and an expanded region extending from the inflow region to the outflow region. The expanded region may have a greater cross-sectional area for fluid flow than each of the inflow region and the outflow region. Objects of the object stream may be moved from the inflow region to the expanded region such that at least a subset of such objects are moved closer to one another. Objects of the object stream may be passed from the expanded region to the outflow region to increase a distance between such objects.

Abstract: Emulsion compositions are provided herein. Also provided herein are kits containing one or more emulsion compositions or components for making such emulsion compositions. Also provided herein are methods of using such emulsion compositions, such as for amplification of target nucleic acids in emulsion droplets.

Abstract: The disclosure provides methods and systems of analyzing single cells by simultaneously separating cells into monodisperse droplets and tagging each nucleic acid molecule from the cells with barcodes unique to each droplet. The methods and systems combine template particles with a plurality of single cells in a tube, generate in the tube monodispersed droplets encapsulating a single one of the template particles and a single one of the single cells, release nucleic acid molecules from the single cells and provide each nucleic acid molecule with a barcode unique to the respective droplet. The nucleic acid molecules can then be analyzed by any known method, for example by sequencing the nucleic acid molecules.

Abstract: The disclosure provides methods and systems of analyzing single cells by simultaneously separating cells into monodisperse droplets and tagging each nucleic acid molecule from the cells with barcodes unique to each droplet. The methods and systems combine template particles with a plurality of single cells in a tube, generate in the tube monodispersed droplets encapsulating a single one of the template particles and a single one of the single cells, release nucleic acid molecules from the single cells and provide each nucleic acid molecule with a barcode unique to the respective droplet. The nucleic acid molecules can then be analyzed by any known method, for example by sequencing the nucleic acid molecules.

Abstract: The invention provides devices that generate monodisperse droplets from a bulk liquid. The devices include a shearing mechanism, a holder for a vessel containing a liquid, and an optical system that transmit light to, and detects light from, liquid in the vessel. The invention also provides methods of using such devices to produce monodisperse droplets.

Abstract: This disclosure provides methods and systems for amplifying small amounts of nucleic acids. Methods include performing amplification reactions in an emulsion format to isolate and clonally amplify discrete populations of nucleic acid molecules inside droplets. In particular, methods and systems of the invention generate an emulsion with particles that template the formation of droplets inside a tube and segregate nucleic acid molecules therein such that each droplet contains an individual nucleic acid molecule. The nucleic acid molecules are amplified inside the droplets.

Abstract: This disclosure provides methods and systems for single-cell analysis, including single-cell transcriptome analysis, of target cells without microfluidic devices. The disclosed methods involve the use of template particles to template the formation of monodisperse droplets to generally capture a single target cell from a population of cells in an encapsulation, derive a plurality of distinct mRNA molecules from the single target cell, and quantify the distinct mRNA molecules to generate an expression profile.

Abstract: Methods to emulsify cells and/or mRNA with reverse transcriptase at a temperature such that the reverse transcriptase begins making cDNA during the emulsification

Abstract: The invention provides methods and systems for drug screening by segregating single cells into droplets simultaneously and providing candidate compound to the single cells to measure cellular response. Methods of the present invention combine template particles with a plurality of single cells in a tube, generate in the tube monodispersed droplets simultaneously that encapsulate a single one of the template particles and single one of the single cells, provide to the single cells one or more candidate compounds, and measure a cellular response to the one or more candidate compounds.

Abstract: Microfluidic systems and methods for arranging a set of objects. In an exemplary method, the set of objects may be transported in carrier fluid along a microfluidic channel structure having a reformatting zone including an object-accessible region and at least one object-excluding region. A portion of the carrier fluid may be moved from the object-accessible region to the at least one object-excluding region in an upstream section of the reformatting zone, to reduce a spacing of objects of the set. The portion of the carrier fluid may be directed into the object-accessible region from the at least one object-excluding region in a downstream section of the reformatting zone, to increase a spacing of objects of the set. The steps of moving and directing in combination may increase the spacing between objects disproportionately for a subset of the objects that are closest to one another.

Abstract: Compositions, devices, and methods are disclosed for the modification of polymer surfaces with coatings having a dispersion of silicone polymer and hydrophobic silica. The surface coatings provide the polymer surface with high hydrophobicity, as well as increased resistance to biofouling with proteinaceous material. The polymer surfaces can be particularly useful in microfluidic devices and methods that involve the contacting of the covalently modified polymer surfaces with emulsions of aqueous droplets containing biological macromolecules within an oil carrier phase.

Abstract: The present disclosure provides a method of hairpin primer design and targeted amplification thereof for creation of targeted sequencing libraries. Generally, the present methods allow for simultaneous construction of targeted sequencing libraries for multiple targeted nucleic acid sequences within a single sample. The presently disclosed methods generate efficient and specific target sequence amplification while avoiding, or significantly reducing, non-specific interaction of multiplex primers, non-specific amplification of sequences due to random priming from molecular “tags” (such as Molecular Identifiers (“MI”) barcodes), and unintentional interactions between gene-specific and universal primers.