An example of a flow cell includes a base support, a reversibly swellable resin positioned over the base support, and a depression defined in the reversibly swellable resin. The reversibly swellable resin includes at least one hydrophilic monomer selected from the group consisting of a poly(ethylene glycol) based monomer, poly(propylene glycol) based monomer, and combinations thereof. The depression has a first opening dimension when the reversibly swellable resin is in a non-swelled stated and has a second opening dimension, that is smaller than the first opening dimension, when the reversibly swellable resin is in a swelled state.
Polynucleotide sequencing methods include incubating unlabeled nucleotides with a cluster of template polynucleotide strands having the same sequence when the identity of the previously added labeled nucleotide is being detected. The detection step provides time for the addition of the unlabeled nucleotides to be incorporated into the copy strands in which the previously added labeled nucleotide did not get incorporated. Thus, at the end of the detection step, all or most of the copy strands will be in phase and ready to incorporate the appropriate labeled nucleotide in the subsequence incorporate step.
C12Q 1/6874 - Methods for sequencing involving nucleic acid arrays, e.g. sequencing by hybridisation [SBH]
B01L 3/00 - Containers or dishes for laboratory use, e.g. laboratory glassware; Droppers
C12Q 1/25 - Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving enzymes not classifiable in groups
3.
Methods of Preparing Directional Tagmentation Sequencing Libraries Using Transposon-Based Technology with Unique Molecular Identifiers for Error Correction
Materials and methods for preparing nucleic acid libraries for next-generation sequencing are described herein. A variety of approaches are described relating to the use of unique molecular identifiers with transposon-based technology in the preparation of sequencing libraries. Also described herein are sequencing materials and methods for identifying and correcting amplification and sequencing errors.
An example nanoimprint lithography (NIL) resin composition includes a total of three monomers, wherein two of the three monomers are selected from the group consisting of two different epoxy substituted silsesquioxane monomers; two different epoxy substituted cyclosiloxane monomers; and two different non-organosilicon epoxy monomers. A third of the three monomers is a fluorinated monomer that is present in an amount ranging from about from 0.5 mass % to about 4 mass %, based on a total solids content of the NIL resin composition. The NIL resin also includes a photoinitiator and a solvent.
C09D 163/00 - Coating compositions based on epoxy resins; Coating compositions based on derivatives of epoxy resins
C08G 59/36 - Epoxy compounds containing three or more epoxy groups together with mono-epoxy compounds
C08G 59/32 - Epoxy compounds containing three or more epoxy groups
C08G 59/68 - Macromolecules obtained by polymerising compounds containing more than one epoxy group per molecule using curing agents or catalysts which react with the epoxy groups characterised by the catalysts used
An example of a sequencing nanoparticle includes a core of a negatively chargeable, hydrophobic polymer. Alternating layers of a positively charged acrylamide hydrogel and the negatively charged polymer are positioned on the core, wherein the positively charged acrylamide hydrogel forms an outer layer of the sequencing nanoparticle. A negatively charged primer set is attached to the outer layer.
A co-polymer includes a plurality of a first monomer including a terminal functional group that is to attach to at least two different primers; a plurality of a second monomer including a second functional group that is different from the terminal functional group, and that is selected from the group consisting of a phenyl group, methoxy propyl, glycosyl, vinyl pyrrolidone, and an imidazole group; and a plurality of a third monomer that is different from the first and second monomers. This co-polymer may be used in a flow cell, and may enhance the clustering efficiency and kinetics.
The present application relates to dyes containing fused tetracyclic bis-boron containing heterocycle and their uses as fluorescent labels. These dyes may be used as fluorescent labels for nucleotides in nucleic acid sequencing applications.
Disclosed herein is a method for enriching a sequencing library comprising double-stranded nucleic acid fragments comprising preparing a library of double-stranded fragments having one or more adaptors at ends of the double-stranded fragment; denaturing the double-stranded fragments to form single-stranded fragments; and hybridizing an extension primer that binds to a target sequence of at least one insert in the library of double-stranded fragments and that does not bind to non-target sequences. In an embodiment, the adaptor is a hairpin adaptor, and extension from the extension primer using a polymerase with 5′ to 3′ exonuclease activity removes all or part of a sequence of the hairpin adaptor that is at least partially complementary to the amplification primer sequence. Each fragment may comprise an insert comprising double-stranded nucleic acid and a hairpin adaptor at the 5′ end of one or both strands of the double-stranded fragments. Hairpin adaptors may comprise an amplification primer sequence and a sequence at least partially complementary to the amplification primer sequence.
In an example method, a series of time-based clustering images is generated for a plurality of library fragments from a genome sample. Each time-based clustering image in the series is sequentially generated. To generate each time-based clustering image in the series: i) a respective sample is introduced to a flow cell, the respective sample including contiguity preserved library fragments of the plurality of library fragments, wherein the contiguity preserved library fragments are attached to a solid support or are attached to each other; ii) the contiguity preserved library fragments are released from the solid support or from each other; iii) the contiguity preserved library fragments are amplified to generate a plurality of respective template strands; iv) the respective template strands are stained; and v) the respective template strands are imaged.
G06F 16/58 - Retrieval characterised by using metadata, e.g. metadata not derived from the content or metadata generated manually
G06V 10/26 - Segmentation of patterns in the image field; Cutting or merging of image elements to establish the pattern region, e.g. clustering-based techniques; Detection of occlusion
B01L 3/00 - Containers or dishes for laboratory use, e.g. laboratory glassware; Droppers
B01L 7/00 - Heating or cooling apparatus; Heat insulating devices
Described herein is a polynucleotide for use as a sequencing template comprising multiple inserts. Also described herein are method of generating and using these polynucleotides and methods of use of such templates, including analysis of contiguity information. Further, sequencing templates comprising an insert sequence and a copy of the insert sequence can be used to correct for random errors generated during sequencing or amplification or to identify nucleobase damage or other mutation that leads to non-canonical base pairing in a double-stranded nucleic acid. Methods of performing methylation analysis are also described herein.
A flow cell includes a support and a heteropolymer attached to the support. The heteropolymer includes an acrylamide monomer including an attachment group to react with a functional group attached to a primer, and a monomer including a stimuli-responsive functional group. The monomer including the stimuli-responsive functional group may be pH-responsive, temperature-responsive, saccharide-responsive, nucleophile-responsive, and/or salt-responsive.
The present disclosure relates to methods, compositions, and kits for treating target nucleic acids, including methods and compositions for fragmenting and tagging nucleic acid (e.g., DNA) using transposome complexes bound to a solid support.
Embodiments of the present disclosure relate to kits, compositions, and methods for nucleic acid sequencing, for example, two-channel nucleic acid sequencing by synthesis using blue and green light excitation. In particular, unlabeled nucleotides for incorporation may be used in conjunction with affinity reagents containing detectable labels excitable by blue and/or green lights, for specific binding to each type of nucleotides incorporated.
The technology disclosed directly operates on sequencing data and derives its own feature filters. It processes a plurality of aligned reads that span a target base position. It combines elegant encoding of the reads with a lightweight analysis to produce good recall and precision using lightweight hardware. For instance, one million training examples of target base variant sites with 50 to 100 reads each can be trained on a single GPU card in less than 10 hours with good recall and precision. A single GPU card is desirable because it a computer with a single GPU is inexpensive, almost universally within reach for users looking at genetic data. It is readily available on could-based platforms.
G16B 40/00 - ICT specially adapted for biostatistics; ICT specially adapted for bioinformatics-related machine learning or data mining, e.g. knowledge discovery or pattern finding
G16B 20/00 - ICT specially adapted for functional genomics or proteomics, e.g. genotype-phenotype associations
G06F 9/38 - Concurrent instruction execution, e.g. pipeline, look ahead
G06N 3/04 - Architecture, e.g. interconnection topology
G06N 3/084 - Backpropagation, e.g. using gradient descent
Provided is a method, including stretching a polynucleotide over a substrate including a plurality of equally spaced cleavage regions including a plurality of transposases, cleaving the polynucleotide with two or more of the plurality of transposases to form a plurality of polynucleotide fragments, and separating, within the plurality of polynucleotide fragments, a population of longer polynucleotide fragments from a population of shorter polynucleotide fragments. Also provided is a method including stretching a polynucleotide over a substrate including a plurality of equally spaced cleavage regions including a plurality of transposases, cleaving the polynucleotide with two or more of the plurality of transposases to form a plurality of polynucleotide fragments, and separating, within the plurality of polynucleotide fragments, a population of longer polynucleotide fragments from a population of shorter polynucleotide fragments.
Reagent cartridges and related systems and methods for controlling reagent temperature are disclosed. In accordance with an implementation, an apparatus includes a system and a reagent cartridge. The system includes a reagent cartridge receptacle, a non-contact temperature controller, a processor operatively coupled to the temperature controller. The reagent cartridge is receivable within the reagent cartridge receptacle and includes a flow cell assembly, a plurality of reagent reservoirs, and a manifold assembly. The manifold assembly includes a common fluidic line and a plurality of reagent fluidic lines. Each of the plurality of reagent fluidic lines is adapted to be fluidically coupled to a corresponding reagent reservoir and selectively couplable to the common fluidic line. The processor is to cause the temperature controller to change a temperature of at least one of the common fluidic line or one or more of the reagent fluidic lines.
Presented herein are altered polymerase enzymes for improved incorporation of nucleotides and nucleotide analogues, in particular altered polymerases that maintain high fidelity under reduced incorporation times, as well as methods and kits using the same.
Embodiments of the present disclosure relate to compositions and kits for use in sequencing by synthesis to improve fluorescent signal intensity and reduce signal decay caused by short wavelength light source during the imaging events. Methods of sequencing using the compositions and kits described herein are also provided.
C07H 21/00 - Compounds containing two or more mononucleotide units having separate phosphate or polyphosphate groups linked by saccharide radicals of nucleoside groups, e.g. nucleic acids
C07H 21/04 - Compounds containing two or more mononucleotide units having separate phosphate or polyphosphate groups linked by saccharide radicals of nucleoside groups, e.g. nucleic acids with deoxyribosyl as saccharide radical
C12Q 1/6809 - Methods for determination or identification of nucleic acids involving differential detection
C12Q 1/6816 - Hybridisation assays characterised by the detection means
20.
METHODS AND SYSTEMS FOR ENCAPSULATING LYOPHILISED MICROSPHERES
The present disclosure relates to a method, including providing one or more lyophilised microspheres in a mixing vessel at a first temperature and generating a fluidized bed of the one or more lyophilised microspheres in the mixing vessel, under conditions effective to encapsulate the one or more lyophilised microspheres with a coating formulation. In an example, the fluidized bed has a fluidization rate of between about 1 cubic meters per hour (m3/h) and about 30 m3/h. In another example, the fluidized bed has an environmental humidity of between about 10% and about 20%. In still another example, the coating formulation is applied at a spray rate of between about 1.5 grams per minute (g/min) and about 10 g/min. In yet another example, the coating formulation is applied at an atomizing rate of between about 0.5 bar and about 1.5 bar. In a further example, the fluidized bed is in a Wurster configuration, a top spray configuration, or a combination thereof. The present disclosure also relates to a system, including one or more lyophilised microspheres, a mixing vessel configured for holding the one or more lyophilised microspheres, a mixer for generating a fluidized bed of the one or more lyophilised microspheres in the mixing vessel at a location, and at least one spray nozzle configured to introduce a shell formulation into the mixing vessel at the location.
The present disclosure relates to methods, compositions, and kits for generating a library of tagged nucleic acid fragments without using PCR amplification, including methods and compositions for fragmenting and tagging nucleic acids (e.g., DNA) using transposome complexes immobilized on solid support.
C12Q 1/6874 - Methods for sequencing involving nucleic acid arrays, e.g. sequencing by hybridisation [SBH]
B01J 19/00 - Chemical, physical or physico-chemical processes in general; Their relevant apparatus
C12Q 1/6806 - Preparing nucleic acids for analysis, e.g. for polymerase chain reaction [PCR] assay
C12Q 1/6837 - Enzymatic or biochemical coupling of nucleic acids to a solid phase using probe arrays or probe chips
C12Q 1/6853 - Nucleic acid amplification reactions using modified primers or templates
C40B 40/06 - Libraries containing nucleotides or polynucleotides, or derivatives thereof
C40B 50/14 - Solid phase synthesis, i.e. wherein one or more library building blocks are bound to a solid support during library creation; Particular methods of cleavage from the solid support
An example of a flow cell includes a substrate; a first primer set attached to a first region on the substrate, the first primer set including an un-cleavable first primer and a cleavable second primer; and a second primer set attached to a second region on the substrate, the second primer set including a cleavable first primer and an un-cleavable second primer.
The present disclosure is concerned with compositions and methods for reducing the steps used in the generation of monoclonal clusters by combining the enzymes used for linearization and removal of unused surface primers.
An example of a flow cell includes a substrate having depressions separated by interstitial regions. First and second primers are immobilized within the depressions. First transposome complexes are immobilized within the depressions, and the first transposome complexes include a first amplification domain. Second transposome complexes are also immobilized within the depressions, and the second transposome complexes include a second amplification domain. Some of the first transposome complexes, or some of the second transposome complexes, or some of both of the first and second transposome complexes include a modification to reduce tagmentation efficiency.
A variety of different types of targeted transposome complexes are described herein that may be used to mediate sequence-specific targeted transposition of nucleic acids. Also described herein is a method of characterizing desired samples in a mixed pool of samples comprising both desired samples and unwanted samples comprising, to produce sequencing data from double-stranded nucleic acid, initially sequencing a library comprising a plurality of nucleic acid samples from a mixed pool, wherein each nucleic acid library comprises nucleic acids from a single sample and a unique sample barcode to distinguish the nucleic acids from the single sample from the nucleic acids from other samples in the library; analyzing the sequencing data and identifying unique sample barcodes associated with sequencing data from desired samples; performing a selection step on the library comprising enriching nucleic acid samples from desired samples and/or depleting nucleic acid samples from unwanted samples; and resequencing the nucleic acid library.
An example of a method includes providing a substrate with an exposed surface comprising a first chemical group, wherein the providing optionally comprises modifying the exposed surface of the substrate to incorporate the first chemical group; reacting the first chemical group with a first reactive group of a functionalized polymer molecule to form a functionalized polymer coating layer covalently bound to the exposed surface of the substrate; grafting a primer to the functionalized polymer coating layer by reacting the primer with a second reactive group of the functionalized polymer coating layer; and forming a water-soluble protective coating on the primer and the functionalized polymer coating layer. Examples of flow cells incorporating examples of the water-soluble protective coating are also disclosed herein.
Embodiments of the present disclosure relate to cyclooctatetraene containing dyes and their uses as fluorescent labels. Also provided are composition containing cyclooctatetraene. The dyes and compositions may be used in various biological applications, such as nucleic acid sequencing.
Embodiments of the present disclosure relate to cyclooctatetraene containing dyes and their uses as fluorescent labels. Also provided are composition containing cyclooctatetraene. The dyes and compositions may be used in various biological applications, such as nucleic acid sequencing.
A polynucleotide sequencing method comprises (i) removing a label and a blocking moiety from a blocked, labeled nucleotide incorporated into a copy polynucleotide strand that is complementary to at least a portion of a template polynucleotide strand; and (ii) washing the removed label and blocking moiety away from the copy strand with a wash solution comprising a first buffer comprising a scavenger compound. Removing the label and blocking moieties may comprise chemically removing the moieties. The first buffer may also comprise an antioxidant and may be used in a scanning buffer used during a nucleotide detection step.
Techniques are described for reducing the number of angles needed in structured illumination imaging of biological samples through the use of patterned flowcells, where nanowells of the patterned flowcells are arranged in, e.g., a square array, or an asymmetrical array. Accordingly, the number of images needed to resolve details of the biological samples is reduced. Techniques are also described for combining structured illumination imaging with line scanning using the patterned flowcells.
The invention relates to methods for pairwise sequencing of a double-stranded polynucleotide template, which permit the sequential determination of nucleotide sequences in two distinct and separate regions on complementary strands of the double-stranded polynucleotide template. The two regions for sequence determination may or may not be complementary to each other.
The present disclosure relates to a method including exposing a composition comprising a wax-microsphere matrix to a first melt-condition, wherein said wax-microsphere matrix comprises a wax component and a plurality of lyophilised microspheres, wherein said plurality of lyophilised microspheres comprise one or more reagent, whereby exposing said composition comprising said wax-microsphere matrix to said first melt-condition melts the wax component; exposing said composition to a first release-condition to rehydrate at least one lyophilised microsphere; and exposing said at least one rehydrated lyophilised microsphere to a separation-condition to separate said wax component from said at least one rehydrated lyophilised microsphere. Also disclosed are methods of preparing a wax-microsphere matrix and releasing one or more reagent from a wax-microsphere matrix as well as compositions. Also disclosed are cartridges with a reagent reservoir including the compositions described herein. Also disclosed are systems for controlling release of one or more reagent including the compositions described herein.
This application describes methods of preparing an immobilized library of tagged RNA fragments. Also described herein are a number of methods of preparing DNA and RNA sequencing libraries from a single sample. These methods can include library preparation from single cells.
The present disclosure relates to new compounds and their use as fluorescent labels. The compounds may be used as fluorescent labels for nucleotides in nucleic acid sequencing applications.
C12Q 1/6816 - Hybridisation assays characterised by the detection means
C09B 23/06 - Methine or polymethine dyes, e.g. cyanine dyes characterised by the methine chain containing an odd number of CH groups three CH groups, e.g. carbocyanines
C07D 209/08 - Indoles; Hydrogenated indoles with only hydrogen atoms or radicals containing only hydrogen and carbon atoms, directly attached to carbon atoms of the hetero ring
C07D 403/06 - Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, not provided for by group containing two hetero rings linked by a carbon chain containing only aliphatic carbon atoms
C09B 23/08 - Methine or polymethine dyes, e.g. cyanine dyes characterised by the methine chain containing an odd number of CH groups more than three CH groups, e.g. polycarbocyanines
G01N 33/58 - Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving labelled substances
C09B 23/10 - Methine or polymethine dyes, e.g. cyanine dyes characterised by the methine chain containing an even number of CH groups
36.
SEQUENCING FROM MULTIPLE PRIMERS TO INCREASE DATA RATE AND DENSITY
The present invention relates to a sequencing method which allows for increased rates of sequencing and an increase in the density of sequencing data. The system may be based on next generation sequencing methods such as sequencing by synthesis (SBS) but uses multiple primers bound at different positions on the same nucleic acid strand.
The present disclosure is concerned with compositions and methods for the paired-end sequencing of target nucleic acids, and more particularly to obtaining nucleotide sequence information from two separate regions of target nucleic acids using amplification sites having a single type of surface primer.
An example of an array includes a support, a cross-linked epoxy polyhedral oligomeric silsesquioxane (POSS) resin film on a surface of the support, and a patterned hydrophobic polymer layer on the cross-linked epoxy POSS resin film. The patterned hydrophobic polymer layer defines exposed discrete areas of the cross-linked epoxy POSS resin film, and a polymer coating is attached to the exposed discrete areas. Another example of an array includes a support, a modified epoxy POSS resin film on a surface of the support, and a patterned hydrophobic polymer layer on the modified epoxy POSS resin film. The modified epoxy POSS resin film includes a polymer growth initiation site, and the patterned hydrophobic polymer layer defines exposed discrete areas of the modified epoxy POSS resin film. A polymer brush is attached to the polymer growth initiation site in the exposed discrete areas.
The invention relates to methods for pairwise sequencing of a polynucleotide template which result in the sequential determination of nucleotide sequence in two distinct and separate regions of the polynucleotide template.
The present disclosure relates to compositions including a shell surrounding an interior compartment, wherein said interior compartment comprises one or more reagent and wherein said shell releases said interior compartment when said shell is exposed to a first release condition, wherein said interior compartment releases said one or more reagent when said interior compartment is exposed to a second release condition, and wherein said first release condition is different from said second release condition. Also disclosed are compositions including a dissolvable first shell, and a dissolvable second shell, the second shell comprising one or more reagent. Also disclosed are methods for controlling release of one or more reagent using the compositions described herein. The present disclosure further relates to cartridges that include a reagent reservoir including the compositions described herein. Also disclosed are systems for controlling release of one or more reagent including the compositions described herein.
Embodiments of the present disclosure relate to nucleotides labeled with photoswitchable compounds. Also provided herein are methods and kits of using these labeled nucleotides for sequencing applications.
Method includes positioning a first carrier assembly on a system stage. The carrier assembly includes a support frame having an inner frame edge that defines a window of the support frame. The first carrier assembly includes a first substrate that is positioned within the window and surrounded by the inner frame edge. The first substrate has a sample thereon. The method includes detecting optical signals from the sample of the first substrate. The method also includes replacing the first carrier assembly on the system stage with a second carrier assembly on the system stage. The second carrier assembly includes the support frame and an adapter plate held by the support frame. The second carrier assembly has a second substrate held by the adapter plate that has a sample thereon. The method also includes detecting optical signals from the sample of the second substrate.
The present application relates to secondary amine-substituted coumarin compounds and their uses as fluorescent labels. The compounds may be used as fluorescent labels for nucleotides in nucleic acid sequencing applications.
C07D 417/04 - Heterocyclic compounds containing two or more hetero rings, at least one ring having nitrogen and sulfur atoms as the only ring hetero atoms, not provided for by group containing two hetero rings directly linked by a ring-member-to-ring- member bond
C07H 19/10 - Pyrimidine radicals with the saccharide radical being esterified by phosphoric or polyphosphoric acids
C12Q 1/68 - Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
C07D 405/04 - Heterocyclic compounds containing both one or more hetero rings having oxygen atoms as the only ring hetero atoms, and one or more rings having nitrogen as the only ring hetero atom containing two hetero rings directly linked by a ring-member-to-ring- member bond
C07D 413/04 - Heterocyclic compounds containing two or more hetero rings, at least one ring having nitrogen and oxygen atoms as the only ring hetero atoms containing two hetero rings directly linked by a ring-member-to-ring- member bond
C12Q 1/6876 - Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes
44.
Exocyclic amine substituted coumarin compounds and uses as fluorescent labels
The present application relates to exocyclic amine-substituted coumarin derivatives and their uses as fluorescent labels. These compounds may be used as fluorescent labels for nucleotides in nucleic acid sequencing applications.
Novel rhodamine dye compounds, labelled conjugates comprising the dyes are described, together with methods for their use. The dyes and labelled conjugates are useful as molecular probes in a variety of applications, such as in assays involving staining of cells, protein binding, and analysis of nucleic acids, such as hybridization assays and nucleic acid sequencing.
An example of a flow cell includes a substrate; a first primer set attached to a first region on the substrate, the first primer set including an un-cleavable first primer and a cleavable second primer; and a second primer set attached to a second region on the substrate, the second primer set including a cleavable first primer and an un-cleavable second primer.
G03F 7/00 - Photomechanical, e.g. photolithographic, production of textured or patterned surfaces, e.g. printed surfaces; Materials therefor, e.g. comprising photoresists; Apparatus specially adapted therefor
The invention relates to methods for pairwise sequencing of a polynucleotide template which result in the sequential determination of nucleotide sequence in two distinct and separate regions of the polynucleotide template.
Presented herein are methods and compositions surface-based tagmentation. In particular embodiments, methods of preparing an immobilized library of fragmented and tagged DNA molecules on a solid surface are presented. In particular embodiments, the solid surface comprises immobilized transposomes in a dried format, suitable for reconstitution upon contact with liquid, such as a liquid sample.
C12Q 1/6806 - Preparing nucleic acids for analysis, e.g. for polymerase chain reaction [PCR] assay
C40B 50/14 - Solid phase synthesis, i.e. wherein one or more library building blocks are bound to a solid support during library creation; Particular methods of cleavage from the solid support
49.
SYSTEMS AND METHODS FOR SEQUENCING NUCLEOTIDES USING TWO OPTICAL CHANNELS
The disclosed technology relates to the field of nucleic acid sequencing, and more particularly, to systems and methods for DNA sequencing utilizing a single optical excitation and at least three fluorescent labels. In some embodiments, the disclosed technology uses a first nucleotide coupled to a first fluorescent label which can emit light to be detectable by a first detector, a second nucleotide coupled to a second fluorescent label which can emit light to be detectable by a second detector, a third nucleotide coupled to a third fluorescent label which can emit light to be detectable by both the first and second detectors, and a fourth nucleotide coupled to no fluorescent label. The disclosed technology may identify a nucleotide in the nucleic acid sequence based on whether the emission is received by the first detector, the second detector, both the first and second detectors, or neither the first nor second detector.
The present application relates to compositions and methods for sequencing by synthesis, where one or more palladium scavengers were used to improve sequencing metrics such phasing and prephasing values.
A method for purifying nucleotides is provided, that includes preparing a solution comprising (a) 3′-blocked nucleotides, (b) 3′-OH nucleotides, (c) a polishing polymerase, and (d) a template. The polishing polymerase and the template are used to selectively polymerize the 3′-OH nucleotides and thus reduce a concentration in the solution of the 3′-OH nucleotides relative to the 3′-blocked nucleotides.
The present application relates to substituted dyes containing bis-boron fused heterocycles and their uses as fluorescent labels. These compounds may be used as fluorescent labels for nucleotides in nucleic acid sequencing applications.
An example of a flow cell includes a substrate and a pattern of two different silanes on at least a portion of a surface of the substrate. A first polymer is attached to a first of the two different silanes and a second polymer is attached to a second of the two different silanes. The first and second polymers respectively include a first functional group and a second functional group of a functional group pair, the functional group pair being selected from the group consisting of an activated ester functional group and an azide functional group, a tetrazine functional group and an activated ester functional group, and a tetrazine functional group and an azide functional group. A first primer set is grafted to the first polymer and a second primer set is grafted to the second polymer. The first and second primer sets are different.
An example method includes introducing a first fluid to a flow channel of a flow cell including a working electrode having a surface that is at least partially exposed to the flow channel, the surface being unmodified or modified with a first member of a transition metal complex binding pair, whereby a linking moiety of a complex present in the first fluid chemically attaches the complex to the surface to form a temporarily modified surface of the working electrode; performing a sensing operation involving the complex of the temporarily modified surface; and applying a desorption voltage of the linking moiety to the working electrode, thereby detaching the linking moiety and regenerating the surface.
A method for detecting different analytes includes mixing different analytes with sensing probes, wherein at least some of the sensing probes are specific to respective ones of the analytes. The analytes respectively are captured by the sensing probes that are specific to those analytes. Fluorophores respectively are coupled to sensing probes that captured respective analytes. The sensing probes are mixed with beads, wherein the beads are specific to respective ones of the sensing probes, and wherein the beads include different codes identifying the analytes to which those sensing probes are specific. The sensing probes respectively are coupled to beads that are specific to those sensing probes. The beads are identified that are coupled to the sensing probes that captured analytes using at least fluorescence from the fluorophores coupled to those sensing probes. The analytes that are captured are identified.
The present disclosure relates to a composition comprising a shell surrounding a core, wherein the core comprises one or more lyophilised microspheres. Also described herein is a method comprising providing one or more lyophilised microspheres; and coating the one or more lyophilised microspheres with a shell under conditions effective to encapsulate the one or more lyophilised microspheres. The present disclosure further relates to a system comprising one or more composition as described herein, and one or more lyophilised cake, wherein the one or more composition and the one or more lyophilised cake are combined under conditions effective to form a rehydration system. Also described herein is a method of controlling release of one or more encapsulated microspheres comprising providing a composition as described herein and mixing the composition with a rehydration solution under a first condition effective to control release of one or more lyophilised microspheres from the composition.
A method for seeding and amplifying target nucleic acids derived from a sample in a cluster at a site on a surface of a substrate includes retaining at least a portion of the target nucleic acids in an inactive form that cannot seed to provide a relatively low concentration of active form target nucleic acids available for seeding. As the active form target nucleic acids seed on the surface of the substrate, they may be amplified. Because the concentration of active form target nucleic acids is low, the likelihood is low that a second active form target nucleic acid will seed at the same site within the same cluster before the first active form target nucleic acid is sufficiently amplified to dominate. Accordingly, the likelihood that the cluster will pass filters is increased relative to traditional seeding and amplification methods employing a higher concentration of active form target nucleic acids.
C12Q 1/6848 - Nucleic acid amplification reactions characterised by the means for preventing contamination or increasing the specificity or sensitivity of an amplification reaction
C12Q 1/70 - Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving virus or bacteriophage
For example, a flowcell includes: a nanowell layer having a first set of nanowells and a second set of nanowells to receive a sample; a first linear waveguide associated with the first set of nanowells, and a second linear waveguide associated with the second set of nanowells; and a first grating for the first linear waveguide, and a second grating for the second linear waveguide, the first and second gratings providing differential coupling of first light and second light.
Examples provided herein are related to detecting methylcytosine and its derivatives using S-adenosyl-L-methionine analogs (xSAMs). Compositions and methods for performing such detection are disclosed. A target polynucleotide may include cytosine (C) and methylcytosine (mC). The method may include (a) protecting the C in the target polynucleotide from deamination; and (b) after step (a), deaminating the mC in the target polynucleotide to form thymine (T). Protecting the C from deamination may include adding a protective group to the 5 position of the C, e.g., using a methyltransferase enzyme that adds the first protective group from an xSAM.
Embodiments provided herein relate to methods and compositions for preparing an immobilized library of barcoded DNA fragments of a target nucleic acid, identifying genomic variants, determining the contiguity information, phasing information, and methylation status of the target nucleic acid.
Provided herein is a method for sequencing a polynucleotide molecules. The method includes the steps of providing a plurality of polynucleotide molecules attached to a surface, wherein a first portion of each polynucleotide molecule is attached to a first location of the surface and a second portion of each polynucleotide molecule is attached to a second location of the surface, the relative proximity of the first and second locations being correlated with the probability that the first and second portions are paired, separating the first and second portions of the polynucleotide molecules on the surface, determining the sequences of the first and second portions of the polynucleotide molecules and comparing the relative proximities and the sequences to determine which first and second portions are paired and to determine the sequence of the target polynucleotide molecules.
A method of generating an asymmetric closed-ended double stranded nucleic acid template from a double stranded nucleic acid template having free 5′ and 3′ ends by use of hairpin or dumbbell adaptors, and sequencing therefrom.
In one example, an unsaturated cyclic dione is coupled to the substrate, and is reacted with an indole or indazole including a first functional group to form a first adduct coupling the first functional group to the substrate. In another example, an unsaturated cyclic dione is coupled to a substrate and reacted with a diene including a functional group to form an adduct coupling the functional group to the substrate. In another example, an indole or indazole is coupled to a substrate, and is reacted with an unsaturated cyclic dione including an oligonucleotide to form an adduct coupling the oligonucleotide to the substrate. In another example, a diene is coupled to a substrate, and is reacted with an unsaturated cyclic dione including an oligonucleotide to form an adduct coupling the oligonucleotide to the substrate.
C08F 283/12 - Macromolecular compounds obtained by polymerising monomers on to polymers provided for in subclass on to polysiloxanes
C07H 21/00 - Compounds containing two or more mononucleotide units having separate phosphate or polyphosphate groups linked by saccharide radicals of nucleoside groups, e.g. nucleic acids
64.
Methods of preparing growing polynucleotides using nucleotides with 3′ AOM blocking group
Embodiments of the present disclosure relate to methods of preparing growing polynucleotides using nucleotide molecules with a 3′ AOM blocking group. Also provided herein are kits related to such methods.
C07H 21/00 - Compounds containing two or more mononucleotide units having separate phosphate or polyphosphate groups linked by saccharide radicals of nucleoside groups, e.g. nucleic acids
65.
LONG STOKES SHIFT CHROMENOQUINOLINE DYES AND USES IN SEQUENCING APPLICATIONS
The present application relates to long Stokes shift chromenoquinoline dyes and their uses as fluorescent labels. For example, these dyes may be used to label nucleotides for nucleic acid sequencing applications.
C12Q 1/6874 - Methods for sequencing involving nucleic acid arrays, e.g. sequencing by hybridisation [SBH]
C07D 491/147 - Ortho-condensed systems the condensed system containing one ring with oxygen as ring hetero atom and two rings with nitrogen as ring hetero atom
C07D 491/22 - Heterocyclic compounds containing in the condensed ring system both one or more rings having oxygen atoms as the only ring hetero atoms and one or more rings having nitrogen atoms as the only ring hetero atoms, not provided for by groups , , or in which the condensed system contains four or more hetero rings
The present disclosure relates to methods, systems, kits and compositions for nucleic acid sequencing applications. In particular, the method utilizes two imaging events with different excitation wavelengths and a single emission channel to collect the fluorescent signal patterns of different types of nucleotide conjugates to determine the identity of the incorporated nucleotide conjugates. The method described herein does not require a chemical treatment of the nucleotide conjugates in the incorporation mixture between the two imaging events.
The present application relates to alkylpyridinium substituted coumarin dyes and their uses as fluorescent labels. For example, these dyes may be used to label nucleotides for nucleic acid sequencing applications.
Embodiments of the present disclosure relate to methods, kits and compositions for two-channel nuclei acid sequencing using blue and violet light excitation (e.g., lasers at 450-460 nm and 400-405 nm respectively). In particular, the nucleotides may be directly labeled with a blue dye, a violet dye, or both a blue dye and a violet dye. Alternatively, one or more nucleotides for incorporation may be unlabeled and affinity reagents containing a blue dye, a violet dye, or both a blue dye and a violet dye may be used to bind specifically to each type of nucleotides incorporated.
Embodiments provided herein relate to methods and compositions for next generation sequencing. Some embodiments include the preparation of a template library from a target nucleic acid in contact with a surface, and sequencing the library on the surface.
Some embodiments of the present application relate to novel modified nucleotide linkers for increasing the efficiency of nucleotide incorporation in Sequencing by Synthesis applications. Methods of preparing these modified nucleotide linkers are also provided herewith.
A hydrogel includes a dendritic core with 2 to 30 arms, and first and second acrylamide monomers incorporated into each arm. The first acrylamide monomer is: (I), wherein R1 and R2 are independently selected from an alkyl, an alkylamino, an alkylamido, an alkylthio, an aryl, a glycol, and optionally substituted variants thereof; and the second acrylamide monomer is: (II), wherein R3 and R4 are independently hydrogen or an alkyl; L is a linker including a linear chain of 2 to 20 atoms selected from carbon, oxygen, and nitrogen and optional substituents on the carbon and any nitrogen atoms; A is an N substituted amide: (III), where R5 is hydrogen or an alkyl; E is a linear chain of 1 to 4 atom(s) selected from carbon, oxygen and nitrogen, and optional substituents on the carbon and any nitrogen atoms; and Z is an optional nitrogen containing heterocycle.
A hydrogel includes a dendritic core with 2 to 30 arms, and first and second acrylamide monomers incorporated into each arm. The first acrylamide monomer is: (I), wherein R1 and R2 are independently selected from an alkyl, an alkylamino, an alkylamido, an alkylthio, an aryl, a glycol, and optionally substituted variants thereof; and the second acrylamide monomer is: (II), wherein R3 and R4 are independently hydrogen or an alkyl; L is a linker including a linear chain of 2 to 20 atoms selected from carbon, oxygen, and nitrogen and optional substituents on the carbon and any nitrogen atoms; A is an N substituted amide: (III), where R5 is hydrogen or an alkyl; E is a linear chain of 1 to 4 atom(s) selected from carbon, oxygen and nitrogen, and optional substituents on the carbon and any nitrogen atoms; and Z is an optional nitrogen containing heterocycle.
An image sensor structure includes an image layer having an array of light detectors disposed therein. A device stack is disposed over the image layer. An array of light guides is disposed in the device stack. Each light guide is associated with a light detector. An array of nanowells is disposed over the device stack. Each nanowell is associated with a first light guide of the array of light guides. A first primer set is disposed throughout a first well region of each nanowell. A second primer set is disposed throughout a second well region of each nanowell. The second well region is adjacent the first well region. The first and second primer sets are operable to attach a forward strand cluster of forward polynucleotide strands in the first well region and a reverse strand cluster of reverse polynucleotide strands in the second well region.
A method for determining the sequence of a target nucleic acid, including steps of contacting a target nucleic acid with a polymerase to sequentially remove nucleotide triphosphates from the target nucleic acid, wherein the nucleotide triphosphates that are removed have a variety of different base moieties; and distinguishing the different base moieties for the nucleotide triphosphates that are removed. Also provided is a apparatus including a nanopore positioned in a fluid impermeable barrier to form a passage through which a nucleotide triphosphate can pass from a first fluid reservoir to a second fluid reservoir, and a reaction mix in the first fluid reservoir that includes a polymerase, target nucleic acid having two strands, and pyrophosphorolytic concentration of pyrophosphate.
The present disclosure provides methods and systems for determining and/or characterizing one or more haplotypes and/or phasing of haplotypes in a nucleic acid sample. In particular, the disclosure provides methods for determining a haplotype and/or phasing of haplotypes in a nucleic acid sample by incorporating synthetic polymorphisms into fragments of a nucleic acid sample and utilizing the synthetic polymorphisms in determining one or more haplotypes and/or phasing of haplotypes.
The present disclosure provides methods and systems for determining and/or characterizing one or more haplotypes and/or phasing of haplotypes in a nucleic acid sample. In particular, the disclosure provides methods for determining a haplotype and/or phasing of haplotypes in a nucleic acid sample by incorporating synthetic polymorphisms into fragments of a nucleic acid sample and utilizing the synthetic polymorphisms in determining one or more haplotypes and/or phasing of haplotypes.
In an example method, a series of time-based clustering images is generated for a plurality of library fragments from a genome sample. Each time-based clustering image in the series is sequentially generated. To generate each time-based clustering image in the series: i) a respective sample is introduced to a flow cell, the respective sample including contiguity preserved library fragments of the plurality of library fragments, wherein the contiguity preserved library fragments are attached to a solid support or are attached to each other; ii) the contiguity preserved library fragments are released from the solid support or from each other; iii) the contiguity preserved library fragments are amplified to generate a plurality of respective template strands; iv) the respective template strands are stained; and v) the respective template strands are imaged.
G06F 16/58 - Retrieval characterised by using metadata, e.g. metadata not derived from the content or metadata generated manually
G06V 10/26 - Segmentation of patterns in the image field; Cutting or merging of image elements to establish the pattern region, e.g. clustering-based techniques; Detection of occlusion
B01L 3/00 - Containers or dishes for laboratory use, e.g. laboratory glassware; Droppers
B01L 7/00 - Heating or cooling apparatus; Heat insulating devices
In an example method, a hydrogel is applied to a surface of a substrate and primers are grafted to the applied hydrogel. Before or after the primers are grafted, plasmonic nanostructures are introduced to the applied hydrogel. This substrate can make up one surface of a flow cell. When the flow cell is used in a sequencing operation, the plasmonic nanostructures can enhance fluorescent signals that are generated.
An example of an incorporation mix includes a liquid carrier, a complex, and a labeled nucleotide. The complex includes a polymerase and a plasmonic nanostructure linked to the polymerase. The labeled nucleotide includes a nucleotide, a 3′ OH blocking group attached to a sugar of the nucleotide, and a dye label attached to a base of the nucleotide.
An example of a functionalized plasmonic nanostructure includes a plasmonic nanostructure core; a polymeric hydrogel attached to the plasmonic nanostructure core, the polymeric hydrogel having a thickness ranging from about 10 nm to about 200 nm; and a plurality of primers attached to side chains or arms of the polymeric hydrogel, wherein at least some of the plurality of primers are attached to the polymeric hydrogel at different distances from the plasmonic nanostructure core.
C12N 15/10 - Processes for the isolation, preparation or purification of DNA or RNA
C08F 265/10 - Macromolecular compounds obtained by polymerising monomers on to polymers of unsaturated monocarboxylic acids or derivatives thereof as defined in group on to polymers of amides or imides
An example of a kit includes a flow cell and a cleavage mix. An example flow cell includes a substrate; a catalytic polymeric hydrogel on the substrate, the catalytic polymeric hydrogel including a deblocking catalyst; and an amplification primer attached to the catalytic polymeric hydrogel. The deblocking catalyst accelerates cleavage of a blocking group of a 3′ OH blocked nucleotide introduced to the flow cell and incorporated into a template strand attached to the amplification primer. An example of the cleavage mix includes a component to initiate cleavage of the blocking group.
C12Q 1/6848 - Nucleic acid amplification reactions characterised by the means for preventing contamination or increasing the specificity or sensitivity of an amplification reaction
A method is used to generate an analysis image of a moving sample based on one or more exposures. An illumination source illuminates a field of view of a camera for one or more pulses while the sample moves through the field of view. The distance moved by the sample during each of these one or more pulses may be less than the size of one pixel in an image captured by the camera.
An analysis substrate comprises: a localization layer to be provided with a sample comprising a nucleotide provided with a fluorescent dye; and a sensor layer comprising an array of sensor pixels, the localization layer being on-chip relative to the sensor layer, one or more of the array of sensor pixels to receive a propagation of fluorescence from the fluorescent dye.
Described herein are methods and systems for performing chemical or enzymatic reactions using electrophoresis. Devices, systems, and methods for preparing a library of tagged nucleic acid fragments from a target double-stranded nucleic acid using electrophoresis are also provided. Application of one or more electric fields causes molecules to migrate through the electrophoresis gel matrix.
For example, a flowcell includes: a nanowell layer having a first set of nanowells and a second set of nanowells to receive a sample; a first linear waveguide associated with the first set of nanowells, and a second linear waveguide associated with the second set of nanowells; and a first grating for the first linear waveguide, and a second grating for the second linear waveguide, the first and second gratings providing differential coupling of first light and second light.
A composition for amplifying a polynucleotide is provided that includes a substrate comprising a first region and a second region. A first plurality of capture primers is coupled to the first region of the substrate. A second plurality of capture primers is coupled to the second region of the substrate. The capture primers of the second plurality of capture primers are longer than the capture primers of the first plurality of capture primers. A first plurality of orthogonal capture primers are coupled to the first region of the substrate. A second plurality of orthogonal capture primers are coupled to the second region of the substrate. The orthogonal capture primers of the second plurality of orthogonal capture primers are shorter than the orthogonal capture primers of the first plurality of orthogonal capture primers.
An imprinting apparatus includes a silicon master having a plurality of nanofeatures defined therein. An anti-stick layer coats the silicon master, the anti-stick layer including a molecule having a cyclosiloxane with at least one silane functional group. A method includes forming a master template by: depositing a formulation on a silicon master including a plurality of nanofeatures defined therein, the formulation including a solvent and a molecule having a cyclosiloxane with at least one silane functional group; and curing the formulation, thereby forming an anti-stick layer on the silicon master, the anti-stick layer including the molecule. The method further includes depositing a silicon-based working stamp material on the anti-stick layer of the master template; curing the silicon-based working stamp material to form a working stamp including a negative replica of the plurality of nanofeatures; and releasing the working stamp from the master template.
B29C 33/38 - SHAPING OR JOINING OF PLASTICS; SHAPING OF MATERIAL IN A PLASTIC STATE, NOT OTHERWISE PROVIDED FOR; AFTER-TREATMENT OF THE SHAPED PRODUCTS, e.g. REPAIRING - Details thereof or accessories therefor characterised by the material or the manufacturing process
C09D 133/26 - Homopolymers or copolymers of acrylamide or methacrylamide
B29C 59/02 - Surface shaping, e.g. embossing; Apparatus therefor by mechanical means, e.g. pressing
G03F 7/00 - Photomechanical, e.g. photolithographic, production of textured or patterned surfaces, e.g. printed surfaces; Materials therefor, e.g. comprising photoresists; Apparatus specially adapted therefor
The present invention relates to a method for preparing a library of template polynucleotides and use thereof in methods of solid-phase nucleic acid amplification. More specifically, the invention relates to a method for preparing a library of template polynucleotides that have common sequences at their 5′ ends and at their 3′ ends.
In a first aspect, a method includes: providing a sample, the sample including a first nucleotide and a second nucleotide; contacting the sample with a first fluorescent dye and a second fluorescent dye, the first fluorescent dye emitting first emitted light within a first wavelength band responsive to a first excitation illumination light, the second fluorescent dye emitting second emitted light within a second wavelength band responsive to a second excitation illumination light; simultaneously collecting, using one or more image detectors, multiplexed fluorescent light comprising the first emitted light and the second emitted light, the first emitted light being a first color channel corresponding to the first wavelength band and the second emitted light being a second color channel corresponding to the second wavelength band; and identifying the first nucleotide based on the first wavelength band of the first color channel and the second nucleotide based on the second wavelength band of the second color channel.
The present invention relates to methods of imaging template hybridisation for estimating cluster numbers prior to solid phase amplification and sequencing. More particularly, an initial round of imaging is carried out at the single molecule template hybridisation stage which allows a general estimation of cluster numbers prior to clusters being formed. Amplification of the signal allows single molecule imaging to be carried out using standard sequencing imaging apparatus.
The assembly includes a docking console and a manifold. The docking console includes a cartridge support surface having a first end and a second end. The manifold has one or more wells defined therein. The docking console further includes a manifold retention bracket to releasably hold the manifold against a fluid cartridge supported on the cartridge support surface at an interface position such that the one or more wells are in fluid communication with the fluid cartridge and a biased seal bar to press the fluid cartridge against the manifold held by the manifold retention bracket. A hydrophilic porous frit disposed within at least one of the wells and is to permit liquid to flow through the outlet aperture but prevent gas from passing through the outlet aperture.
The present application relates to substituted coumarin derivatives and their uses as fluorescent labels. These compounds may be used as fluorescent labels for nucleotides in nucleic acid sequencing applications.
C12Q 1/6876 - Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes
C07D 407/04 - Heterocyclic compounds containing two or more hetero rings, at least one ring having oxygen atoms as the only ring hetero atoms, not provided for by group containing two hetero rings directly linked by a ring-member-to-ring- member bond
C07D 409/04 - Heterocyclic compounds containing two or more hetero rings, at least one ring having sulfur atoms as the only ring hetero atoms containing two hetero rings directly linked by a ring-member-to-ring- member bond
C07D 405/14 - Heterocyclic compounds containing both one or more hetero rings having oxygen atoms as the only ring hetero atoms, and one or more rings having nitrogen as the only ring hetero atom containing three or more hetero rings
C07D 409/14 - Heterocyclic compounds containing two or more hetero rings, at least one ring having sulfur atoms as the only ring hetero atoms containing three or more hetero rings
C07D 417/04 - Heterocyclic compounds containing two or more hetero rings, at least one ring having nitrogen and sulfur atoms as the only ring hetero atoms, not provided for by group containing two hetero rings directly linked by a ring-member-to-ring- member bond
C07D 417/14 - Heterocyclic compounds containing two or more hetero rings, at least one ring having nitrogen and sulfur atoms as the only ring hetero atoms, not provided for by group containing three or more hetero rings
C07D 413/04 - Heterocyclic compounds containing two or more hetero rings, at least one ring having nitrogen and oxygen atoms as the only ring hetero atoms containing two hetero rings directly linked by a ring-member-to-ring- member bond
The invention relates to modified polymerase enzymes which exhibit improved incorporation of nucleotide analogues bearing substituents at the 3′ position of the sugar moiety that are larger in size than the naturally occurring 3′hydroxyl group. Also described are methods of using the polymerases to incorporate nucleotides into polynucleotides, particularly in the context of DNA sequencing.
Provided is a method, including stretching a polynucleotide over a substrate including a plurality of equally spaced cleavage regions including a plurality of transposases, cleaving the polynucleotide with two or more of the plurality of transposases to form a plurality of polynucleotide fragments, and separating, within the plurality of polynucleotide fragments, a population of longer polynucleotide fragments from a population of shorter polynucleotide fragments. Also provided is a method including stretching a polynucleotide over a substrate including a plurality of equally spaced cleavage regions including a plurality of transposases, cleaving the polynucleotide with two or more of the plurality of transposases to form a plurality of polynucleotide fragments, and separating, within the plurality of polynucleotide fragments, a population of longer polynucleotide fragments from a population of shorter polynucleotide fragments.
Presented are methods and compositions for preparing samples for amplification and sequencing. Particular embodiments relate to methods of preparing nucleic acid-containing cellular samples for library amplification, wherein the methods include lysing cells of the sample to form a lysate, amplifying the nucleic acids from the lysed samples, exposing the amplified nucleic acids to a solid surface, and clonallyr amplifying the amplified nucleic acids to generate clusters.
Embodiments of the present disclosure relate to nucleotide and nucleoside molecules with acetal 3′-OH blocking groups. Also provided herein are methods to prepare such nucleotide and nucleoside molecules, and the uses of fully functionalized nucleotides containing the 3′ acetal blocking group for sequencing applications.
The present disclosure is concerned with compositions, methods, and kits for preparing a sequencing library. In one embodiment, methods include producing a library of target nucleic acids having the same adapter at each end and then switching the identity of one adapter to result in target nucleic acids flanked by distinct adapters.
Techniques are described for reducing the number of angles needed in structured illumination imaging of biological samples through the use of patterned flowcells, where nanowells of the patterned flowcells are arranged in, e.g., a square array, or an asymmetrical array. Accordingly, the number of images needed to resolve details of the biological samples is reduced. Techniques are also described for combining structured illumination imaging with line scanning using the patterned flowcells.
A method of sequencing a plurality of polynucleotides includes: attaching a single DNA template molecule to each of a plurality of attachment elements on a sample container, wherein the average distance between adjacent elements is less than Abbe's limit; applying a stochastic photo-switching chemistry to all of the molecules at the same time to cause the attached molecules to fluoresce in on and off events in up to four different colors by stochastic photo-switching; and imaging the on and off events in a color channel for each color in real-time as the on and off events are occurring for the attached molecules.
Presented herein are altered polymerase enzymes for improved incorporation of nucleotides and nucleotide analogues, in particular altered polymerases that maintain high fidelity under reduced incorporation times, as well as methods and kits using the same.
An example of a flow cell includes a substrate and a cured, patterned resin on the substrate. The cured, patterned resin has nano-depressions separated by interstitial regions. Each nano-depression has a largest opening dimension ranging from about 10 nm to about 1000 nm. The cured, patterned resin also includes an interpenetrating polymer network. The interpenetrating polymer network of the cured, patterned resin includes an epoxy-based polymer and a (meth)acryloyl-based polymer.
C08G 59/06 - Polycondensates containing more than one epoxy group per molecule of polyhydroxy compounds with epihalohydrins or precursors thereof of polyhydric phenols
C08G 59/32 - Epoxy compounds containing three or more epoxy groups
B01L 3/00 - Containers or dishes for laboratory use, e.g. laboratory glassware; Droppers