Aptamer detection techniques with dynamic range compression are described that permit removal of a portion of more abundant aptamers in an aptamer-based assay. In an embodiment, a mixture of tagged probes and dummy probes can be used such that the dummy probes bind abundant aptamers and in turn are not captured or amplified for detection in downstream steps. Other techniques are also contemplated, including targeted removal of or cleavage of probes that bind to excess aptamers.
The present disclosure is concerned with proteins, methods, compositions, and kits for mapping of methylation status of nucleic acids, including 5-methylcytosine and 5-hydroxymethyl cytosine (5hmC). In one embodiment, proteins are provided that selectively act on certain modified cytosines of target nucleic acids and converts them to thymidine or modified thymidine analogues. In another embodiment, proteins are provided that selectively act on certain modified cytosines of target nucleic acids and converts them to uracil or thymidine and selectively do not act on other certain modified cytosines of target nucleic acids. Also provided are compositions and kits that include one or more of the proteins and methods for using one or more of the proteins.
Embodiments of the present disclosure relate to modified extension primers for use in generating clustered polynucleotides for sequencing by synthesis. In particular, the disclosure relates to methods of chemically linearizing clustered polynucleotides in preparation for sequencing by cleavage of one or more strands of double-stranded polynucleotides immobilized on a solid support by a periodate salt.
Embodiments of the present disclosure relate to compositions and methods for improving the intensity of the fluorescent signals during nucleic acid sequencing. In particular, at least one biotin-binding site of the labeled streptavidin is blocked to reduce fluorescent signal deflation.
The present application relates to chromenoquinoline dyes and their uses as fluorescent labels. For example, these dyes may be used to label nucleotides for nucleic acid sequencing.
C07D 491/052 - Ortho-condensed systems with only one oxygen atom as ring hetero atom in the oxygen-containing ring the oxygen-containing ring being six-membered
A61K 31/7105 - Natural ribonucleic acids, i.e. containing only riboses attached to adenine, guanine, cytosine or uracil and having 3'-5' phosphodiester links
A61K 31/711 - Natural deoxyribonucleic acids, i.e. containing only 2'-deoxyriboses attached to adenine, guanine, cytosine or thymine and having 3'-5' phosphodiester links
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
C07H 19/10 - Pyrimidine radicals with the saccharide radical being esterified by phosphoric or polyphosphoric acids
C07H 19/20 - Purine radicals with the saccharide radical being esterified by phosphoric or polyphosphoric acids
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
6.
COMPOSITIONS AND METHODS FOR IMPROVING SEQUENCING SIGNALS
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.
Library preparation systems and methods are disclosed. An apparatus includes a plate receptacle, a magnet, a thermocycler, and an actuator. The plate receptacle is to receive a plate having a well and the thermocycler is to adjust a temperature of a sample within the well of the plate and the actuator is to move the magnet relative to the plate receptacle.
B01L 3/00 - Containers or dishes for laboratory use, e.g. laboratory glasswareDroppers
C40B 50/14 - Solid phase synthesis, i.e. wherein one or more library building blocks are bound to a solid support during library creationParticular methods of cleavage from the solid support
C40B 60/14 - Apparatus specially adapted for use in combinatorial chemistry or with libraries for creating libraries
8.
NUCLEOSIDES AND NUCLEOTIDES WITH 3' VINYL BLOCKING GROUP
Embodiments of the present disclosure relate to nucleotide and nucleoside molecules with an optionally substituted 3'vinyl blocking group. Also provided herein are methods to prepare such nucleotide and nucleoside molecules. Additionally, the present disclosure provides methods of using such blocked nucleosides and nucleotides in oligonucleotide synthesis and sequencing.
The present disclosure is generally directed to strategies for template capture and amplification during sequencing. In some examples, a solid support is used for template capture and amplification.
Embodiments of the present disclosure also relate to methods of fabricating flow cell substrates. Some exemplary workflows exploit orthogonal chemistries of substrate layers such that the process does not include polishing steps. Substrates prepared by the method described herein can include a first primer set and a second primer set compatible with simultaneous paired-end sequencing methods.
G03F 7/00 - Photomechanical, e.g. photolithographic, production of textured or patterned surfaces, e.g. printed surfacesMaterials therefor, e.g. comprising photoresistsApparatus specially adapted therefor
A sequencing kit includes a plurality of particles and a flow cell. The plurality of particles includes a primer set attached to a surface of each of the plurality of particles; and a flow cell surface attachment mechanism attached to the surface of each of the plurality of particles. The flow cell surface attachment mechanism is selected from the group consisting of a capture primer, an alkene, an alkyne, biotin, and a charged polymer. The flow cell includes a plurality of chemical pads that are spatially separated from one another on a substantially flat substrate surface, each of the chemical pads including chemistry to attach to the surface attachment mechanism.
Reagent cartridges and related systems and methods are disclosed. In accordance with an implementation, an apparatus includes a first flexible container, a second flexible container, and a coupling. The first flexible container has an end and defines a first interior containing reagent. The second flexible container has an end and defines a second interior. The first flexible container is positioned within the second interior. The coupling has a first portion coupled to the end of the first flexible container and a second portion coupled to the end of the second flexible container. The coupling has a reagent port fluidly coupled to the first interior of the first flexible container and a pressure port fluidly coupled to the second interior of the second flexible container.
The invention relates to methods and associated products for preparing polynucleotide sequences for detection of modified cytosines and sequencing said polynucleotides to detect modified cytosines. The methods comprise treatment of the target polynucleotide with a conversion reagent that is configured to convert a modified cytosine to thymine or a nucleobase which is read as thymine/uracil, and/or configured to convert an unmodified cytosine to uracil or a nucleobase which is read as thymine/uracil. In particular embodiments, portions of both strands of the treated target are sequenced concurrently.
Disclosed herein include systems, machines, devices, and methods for single-pass methylation mapping. C-to-T converted sequence reads and G-to-A converted sequence reads generated from a sample subjected to a methylation assay can be mapped to a mapping reference sequence comprising a C-to-T converted reference sequence and a G-to-A converted reference sequence generated to a reference genome sequence. The counts of Cs and Ts of sequence reads mapped to each of one or more positions with Cs in the reference genome sequence can be used to determine whether the position is a methylated C or an unmethylated C in the sample.
Non-contact dispensers and related methods are disclosed. In an implementation, an apparatus includes a plate receptacle, a reagent reservoir receptacle, and a non-contact dispenser. The reagent reservoir receptacle receives a reagent reservoir containing reagent and the non-contact dispenser includes an inlet, an upstream valve, a syringe pump, a downstream valve, and an outlet. The syringe pump has a barrel and a hollow plunger movably disposed within the barrel. A flow path is defined between the inlet and the outlet and through the upstream valve, the barrel, the hollow plunger, and the downstream valve. The barrel is fluidically between the upstream valve and the downstream valve. The plate receptacle is to receive a well plate having a well and the non-contact dispenser is to dispense the reagent from the reagent reservoir into the well of the plate.
B67D 7/02 - Apparatus or devices for transferring liquids from bulk storage containers or reservoirs into vehicles or into portable containers, e.g. for retail sale purposes for transferring liquids other than fuel or lubricants
B67D 7/36 - Arrangements of flow- or pressure-control valves
17.
FUNCTIONALIZED NANOSTRUCTURES AND FLOW CELL DEPRESSIONS
A functionalized nanostructure includes a metal nanostructure; an un-cleavable first primer and a cleavable second primer attached to a first region of the metal nanostructure through i) a first thiol linkage attached to a first polymer chain having a first polarity or ii) respective first thiol linkages attached to respective first polymer chains having the first polarity; and a cleavable first primer and an un-cleavable second primer attached to a second region of the metal nanostructure through i) a second thiol linkage attached to a second polymer chain having a second polarity different from the first polarity or ii) respective second thiol linkages attached to respective second polymer chains having the second polarity.
This disclosure describes methods, non-transitory computer readable media, and systems that can use a machine-learning to determine factors or scores indicating an error level with which a given methylation assay detects methylation of cytosine bases. For instance, the disclosed systems use a machine-learning model to generate a bias score indicating a degree to which a given methylation assay errs in detecting cytosine methylation when specific sequence contexts surround such cytosines compared to other sequence contexts. The machine-learning model may take various forms of models, including a decision-tree model, a neural network, or a combination of a decision-tree model and a neural network. In some cases, the disclosed system combines or uses bias scores from multiple machine-learning models to generate a consensus bias score.
G16B 20/20 - Allele or variant detection, e.g. single nucleotide polymorphism [SNP] detection
G16B 40/00 - ICT specially adapted for biostatisticsICT specially adapted for bioinformatics-related machine learning or data mining, e.g. knowledge discovery or pattern finding
19.
MINIMIZING FETAL FRACTION BIAS IN MATERNAL POLYGENIC RISK SCORE ESTIMATION
The presently described techniques provide for the use of low-pass sequencing data in the calculation of a polygenic risk score (14) for an individual. As discussed herein, the low-pass sequencing data may be acquired in a context where DNA (e.g., cfDNA (350)) from more than one source is present in the sample and the portion of the DNA attributable to a secondary source may bias the PRS calculation for the primary individual of interest. In one implementation fragment length may be used to derive a function (e.g., a linear function) relating fetal fraction to the respective PRS estimate at each fetal fraction. This function may then be used to calculate the PRS in the absence of a fetal contribution (i.e., at a 0% fetal fraction).
G16B 20/20 - Allele or variant detection, e.g. single nucleotide polymorphism [SNP] detection
G16H 50/20 - ICT specially adapted for medical diagnosis, medical simulation or medical data miningICT specially adapted for detecting, monitoring or modelling epidemics or pandemics for computer-aided diagnosis, e.g. based on medical expert systems
G16B 40/00 - ICT specially adapted for biostatisticsICT specially adapted for bioinformatics-related machine learning or data mining, e.g. knowledge discovery or pattern finding
Some implementations of the disclosure relate to an imaging system, including: a sample holder to support a sample container having multiple sample locations; an optical stage having; an assembly comprising one or more actuators physically coupled to the sample holder to tilt the sample holder relative to the optical stage during imaging of the multiple sample locations to focus the optical stage onto a current sample location; a first light source to project a first pair of spots on the sample container; and a controller to control, based on a sample tilt determined from a first separation measurement of the first pair of spots from one or more images taken by an image sensor at one or more of the sample locations, the one or more actuators to tilt the sample holder along a first direction of the imaging or a second direction substantially perpendicular to the first direction.
Embodiments of the present disclosure relates to various bisulfite-free chemical methods for detecting methylation of cytosine in the DNA sample. These methods convert methylated and hydroxymethylated cytosine in the nucleic acid sequence to a modified or pseudo thymine or a uracil moiety which then can be detected in sequencing.
Polynucleotide sequencing methods for sequencing one or more polynucleotide templates that uses primers bound to a surface as sequencing primers. The surface primers may include at least a portion of a surface oligonucleotide used during cluster formation. The sequencing methods may be used for single stranded sequencing or double stranded sequencing. Double stranded sequencing methods may employ an enzyme that has nick-translation activity. A kit includes all the reagents needed for sequencing does not include sequencing primers. The kit may be used to accomplish the sequencing methods of the present disclosure.
Imaging systems and related methods are disclosed. In accordance with an implementation, a system includes a flow cell receptacle to receive a flow cell that receives a sample and an imaging system having a light source assembly, and an imaging device. The light source assembly to form a substantially collimated beam. The optical assembly including an asymmetric beam expander group that includes one or more asymmetric elements or anamorphic elements disposed along an optical axis. The optical assembly to receive the substantially collimated beam from the light source assembly, and transform the substantially collimated beam into a shaped sampling beam having an elongated cross section in a far field at or near a focal plane of the optical assembly to optically probe the sample. The imaging device to obtain image data associated with the sample in response to the optical probing of the sample with the sampling beam.
This disclosure describes methods, non-transitory computer readable media, and systems that can flexibly and efficiently change versions of a variant analysis model for different genomic analysis applications. For example, the disclosed systems can determine a particular version of a variant analysis model indicated by a genomic analysis application and can update a genomic analysis device (e.g., FPGA, CPU) by installing the indicated version of the variant analysis model. The disclosed systems can further execute a genomic analysis application to analyze nucleotide base calls utilizing the version of variant analysis model indicated by the genomic analysis application.
G16B 20/20 - Allele or variant detection, e.g. single nucleotide polymorphism [SNP] detection
G16B 5/00 - ICT specially adapted for modelling or simulations in systems biology, e.g. gene-regulatory networks, protein interaction networks or metabolic networks
A computer-implemented method of quantifying a strength of association of genes associated with a phenotype and a contribution of rare variants to a phenotype response by calculating a weighted burden score (162) for a plurality of associated genes with a specified phenotype, wherein the burden score (162) identifies identifying consequential, non-random association in a cohort (104) between carrier status of each of the associated genes and a phenotype response to presence in the associated genes of one or more rare pathogenic variants. Respective effective strength scores are determined (142) for the consequential, non-random association for genes selected from the associated genes based on respective burden scores (162) at per-gene resolution.
Embodiments of the present disclosure relates to periodate salt compositions for use in the chemical linearization of double-stranded polynucleotides in preparation for sequencing application, for example, sequencing-by-synthesis (SBS). Kits containing the periodate salt composition and methods of sequencing polynucleotides are also described.
Embodiments of the present disclosure relates to periodate salt compositions for use in the chemical linearization of double-stranded polynucleotides in preparation for sequencing application, for example, sequencing-by-synthesis (SBS). Kits containing the periodate salt composition and methods of sequencing polynucleotides are also described.
Systems and related temperature calibration methods. In accordance with a first implementation, an apparatus includes a flow cell interface, a temperature control device, an infrared sensor, and a controller. The flow cell interface includes a flow cell support and the temperature control device is for the flow cell support. The controller is to command the temperature control device to cause the flow cell support to achieve a temperature value, cause the infrared sensor to measure an actual temperature value of the flow cell support, and calibrate the temperature control device based on a difference between the commanded temperature value and the actual temperature value.
G01J 5/061 - Arrangements for eliminating effects of disturbing radiationArrangements for compensating changes in sensitivity by controlling the temperature of the apparatus or parts thereof, e.g. using cooling means or thermostats
29.
INSTRUMENTS, SYSTEMS, AND METHODS FOR MEASURING LIQUID FLOW THROUGH CHANNELS
In some examples, an instrument for measuring a volume of a liquid is provided. A gas flow rate sensor may measure a rate of flow of a pressurized gas to a reservoir storing a liquid. A controller may be coupled to the gas flow rate sensor and may calculate a volume of the liquid that the flow of pressurized gas displaces from the reservoir. In some examples, a method of measuring a volume of a liquid is provided. Using a gas flow rate sensor, a flow of pressurized gas may be measured. The flow of the pressurized gas may be delivered to a reservoir storing a liquid. A volume of the liquid in the reservoir may be displaced using the flow of pressurized gas. The measurement of the flow of the pressurized gas may be used to calculate the volume of the liquid that is displaced.
G01F 15/00 - Details of, or accessories for, apparatus of groups insofar as such details or appliances are not adapted to particular types of such apparatus
G01F 15/07 - Integration to give total flow, e.g. using mechanically-operated integrating mechanism
G01F 23/22 - Indicating or measuring liquid level or level of fluent solid material, e.g. indicating in terms of volume or indicating by means of an alarm by measuring physical variables, other than linear dimensions, pressure or weight, dependent on the level to be measured, e.g. by difference of heat transfer of steam or water
Embodiments of the present disclosure relate to method of chemical linearization of double stranded polynucleotides for sequencing by synthesis. In particular, a heterogenous cobalt catalyst is used to cleave one or more diol moieties at a predetermined cleavage site of one strand of the double stranded polynucleotides.
The present disclosure is directed to decoupling library capture (template seeding) from cluster generation to optimise both processes. This is achieved by introducing orthogonality between the seeding and clustering primer.
Reagent reservoirs and related systems and methods are disclosed. In accordance with a first implementation, an apparatus includes a system and a reagent reservoir. The system includes a reagent reservoir receptacle. The reagent reservoir is received within the reagent reservoir receptacle and has a body and a fluidic port. The body defines a storage chamber, a sipper chamber, and a fluidic sinus fluidly coupling the storage chamber and the sipper chamber. The fluidic port is fluidly coupled to the sipper chamber.
Examples herein include an apparatus having a substrate, a sensor over the substrate including an active surface and a sensor bond pad, a molding layer over the substrate and covering sides of the sensor, the molding layer having a lower portion with a first molding height relative to a top surface of the substrate and an upper portion with a second molding height relative to the top surface of the substrate, the first molding height and the second molding height each being greater than a height of the active surface, the second molding height being great than the first molding height; and a lidding layer over at least some of the lower portion of the molding layer and over the active surface. The lidding layer and the molding layer form a space over the active surface of the sensor that defines a flow channel.
An example primer set includes first and second nuclease resistant primers. The first nuclease resistant primer includes a first sequence; a first cleavage site attached at a 3' end of the first sequence; and a first nuclease resistant modification incorporated between the first sequence and the first cleavage site. The second nuclease resistant primer includes a second sequence that is different from the first sequence; a second nuclease resistant modification incorporated at a 3' end of the second sequence; and a second cleavage site attached between the second sequence and the second nuclease resistant modification. The second cleavage site is different from the first cleavage site.
Systems and related manifold assemblies are disclosed. In accordance with an implementation, an apparatus comprises or includes a cassette assembly of a sipper manifold assembly comprising or having a cassette housing, one or more sipper tubes, one or more sipper couplings, and one or more biasing elements. The one or more sipper tubes comprise or have a proximal end and a distal end. The one or more sipper couplings is movably coupled to the cassette housing the proximal end of the sipper tubes are coupled to the sipper couplings. The one or more biasing elements bias the one or more sipper couplings. The one or more biasing elements allow relative movement between the sipper tubes and the cassette housing.
Embodiments of the present disclosure relate to methods of capturing library DNA complexes to the patterned surface of the solid support for sequencing. The methods described herein improve the monoclonality of clusters and sequencing data quality and read length.
A polynucleotide sequencing method includes a wash step that employs a composition including a polymerase. The composition may also include a plurality of nucleotides. The composition may be configured to prevent the polymerase from incorporating one of the plurality of nucleotides into a copy polynucleotide strand. The composition may be substantially free of Mg2.
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 1/6874 - Methods for sequencing involving nucleic acid arrays, e.g. sequencing by hybridisation [SBH]
40.
SOLID SUPPORTS AND METHODS FOR DEPLETING AND/OR ENRICHING LIBRARY FRAGMENTS PREPARED FROM BIOSAMPLES
Described herein are solid supports and methods for depleting library fragments prepared from unwanted RNA sequences and/or enriching library fragments prepared from desired RNA sequences. These methods may incorporate microfluidics and flow cells for greater ease of use. Libraries enriched or depleted with the present methods may be used for sequencing, e.g. by bridge sequencing. Also described are probes specific for unwanted RNA and methods for enzymatic depletion, using RNAse H, of ribosomal RNA from human microbiome samples, whereby DNAse I may be used for degrading the probes and magnetic beads may be used for binding the desired RNA.
An example flow cell includes a substrate having a surface. The flow cell also includes a polymeric hydrogel attached to at least a portion of the substrate surface, where the polymeric hydrogel includes a dark quencher. The flow cell further includes at least one primer set attached to the polymeric hydrogel.
The invention relates to methods of preventing renaturation of single-stranded nucleic acid libraries during storage, the method comprising using blocking oligonucleotides substantially complementary to adaptor sequences in the nucleic acid library.
Systems and methods for sequencing polynucleotides using nanopores are disclosed. In some embodiments, a polynucleotide including a single-stranded region and a double-stranded region, in which the single-stranded region is disposed through a nanopore. The polynucleotide can be moved relative to the nanopore by electric forces while one or more structural locks keep the polynucleotide close to the nanopore. A characteristic signal based on nanopore ionic current blockade and associated with the regions of the polynucleotide at or near the nanopore recognition zone is measured and used to infer the nucleobase sequence of the polynucleotide. In some examples, the double-stranded region is extended by a polymerase, and the polymerase is removed from the polynucleotide. In some examples, signals measured under different applied voltages provide nonredundant information regarding the polynucleotide sequence.
Sequencing polynucleotides using nanopores is provided herein. A polynucleotide is disposed through a nanopore's aperture such that its 3' end is on the nanopore's first side and its 5' end is on the nanopore's second side. On the nanopore's first side, a duplex with the polynucleotide is formed that includes a 3' end. The duplex is extended on the first side of the nanopore by adding a nucleotide to the 3' end of the duplex. A first force is applied disposing the 3' end of the duplex within the aperture, and the nanopore inhibits translocation of the 3' end of the duplex to the second side of the nanopore. A value of an electrical property of the 3' end of the duplex and a single-stranded portion of the polynucleotide is measured. The nucleotide at the 3' end of the duplex is identified using the measured value.
Some of the resin compositions are ultraviolet light or thermally curable, while others are ultraviolet light curable. One example of the ultraviolet light or thermally curable resin composition consists of a predetermined mass ratio of a (meth)acrylate cyclosiloxane monomer and a non-siloxane (meth)acrylate based monomer ranging from about >0:<100 to about 80:20; from 0 mass% to about 10 mass%, based on a total solids content of the resin composition, of an initiator selected from the group consisting of an azo-initiator, an acetophenone, a phosphine oxide, a brominated aromatic acrylate, and a dithiocarbamate; a surface additive; and a solvent.
An example of an ultraviolet light curable resin composition includes a predetermined mass ratio of a first epoxy substituted polyhedral oligomeric silsesquioxane monomer and a second substituted polyhedral oligomeric silsesquioxane monomer, wherein the first and second epoxy substituted polyhedral oligomeric silsesquioxane monomers are different, and wherein the predetermined mass ratio ranges from about 3:7 to about 7:3; bis-(4-methylphenyl)iodonium hexafluorophosphate as a first initiator; a second initiator selected from the group consisting of a free radical initiator and a cationic initiator other than bis-(4- methylphenyl)iodonium hexafluorophosphate; a surface additive; and a solvent.
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.
Generation and use of a focus quality metric that is intensity independent is described. In one example, the focus quality metric is generated by acquiring an image, such as an image of a patterned surface of a flow cell, and processing all or part (e.g., a sub-region or sub- image) to generate a Fourier transform of the respective image data. By way of example, in one embodiment a discrete Fourier transform may be applied to a sub-region of an image of a patterned flow cell surface. A focus quality metric that is intensity independent may be derived from the Fourier transform of the image data.
An apparatus includes a flow cell body with an array of reaction sites positioned along a floor of a channel. An optical filter layer is positioned under the floor of the channel and includes at least a portion spanning uninterruptedly along a length corresponding to the length of the array of reaction sites. Imaging regions are positioned under the optical filter layer. Each imaging region is positioned directly under a corresponding reaction site. The optical filter layer is configured to permit one or more selected wavelengths of light to pass from each reaction site to the imaging region forming a sensing pair with the reaction site. The optical filter layer is configured to reduce transmission of excitation light directed toward the reaction sites; and to reduce transmission of light emitted from each reaction site to imaging regions not forming a sensing pair with the reaction site.
Disclosed herein include methods, compositions, reaction mixtures, kits and systems for identification of methylated cytosines in nucleic acids using a bisulfite-free, one-step chemoenzymatic modification of methylated cytosines.
Presented herein are altered polymerase enzymes for improved incorporation of nucleotides and nucleotide analogues, in particular altered polymerases that maintain low pre-phasing rates when using ambiently stored polymerases, as well as methods and kits using the same.
Detecting analytes using proximity-induced tagmentation, strand invasion, restriction, or ligation is provided herein. In some examples, detecting an analyte includes coupling a donor recognition probe to a first portion of the analyte. The donor recognition probe includes a first recognition element specific to the first portion of the analyte, a first oligonucleotide corresponding to the first portion, and a transposase coupled to the first recognition element and the first oligonucleotide. An acceptor recognition probe is coupled to a second portion of the analyte. The acceptor recognition probe includes a second recognition element specific to the second portion of the analyte and a second oligonucleotide coupled to the second recognition element and corresponding to the second portion. The transposase is used to generate a reporter polynucleotide including the first and second oligonucleotides. The analyte is detected based on the reporter including comprising the first and second oligonucleotides.
Some embodiments relate to the preparation of nucleic acid libraries for detecting genomic methylation. Some embodiments include the use of hairpin adapters to physically link a conversion-sensitive strand with a conversion-resistant strand. Some embodiments include the use of adapters comprising tags such that a sequence derived from a template strand can be matched with a sequence derived from the complementary strand of the nucleic acid of the sample.
A method of generating base calls by a base caller is disclosed. The method includes receiving a plurality of sensor data from a flow cell, wherein the plurality of sensor data is within a first range and identifying a second range, such that at least a threshold percentage of the plurality of sensor data are within the second range. At least a subset of the plurality of sensor data, that are within the second range, are mapped to a third range, thereby generating a plurality of normalized sensor data. The plurality of normalized sensor data is processed in a base caller, to call, for the plurality of normalized sensor data, one or more corresponding bases.
Devices for sequencing biopolymers, methods of manufacturing the devices, and methods of using the devices are disclosed. In one example, such a device has a nanopore and a horizontal nanochannel. In some embodiments, the horizontal nanochannel may take a tortuous path. In some embodiments, such a device includes gas or air bubble generators or pressure pulse generators to block or unblock the horizontal nanochannel.
Techniques for improving artificial intelligence-based base calling are disclosed. The improved techniques can be used to better train artificial intelligence for base calling by reordering of sequencing images, and training of a neural network-based base caller where the temporal logic is effectively "frozen" (or bypassed). In addition, the improved techniques include various combinations, including, for example, combining "normalization" of sequencing images with reordering of sequencing images and/or with effectively "freezing" the temporal logic.
A method of progressively training a base caller is disclosed. The method includes iteratively initially training a base caller with analyte comprising a single-oligo base sequence, and generating labelled training data using the initially trained base caller. At operations (i), the base caller is further trained with analyte comprising multi-oligo base sequences, and labelled training data is generated using the further trained base caller. Operations (i) are iteratively repeated to further train the base caller. In an example, during at least one iteration, a complexity of neural network configuration loaded within the base caller is increased. In an example, labelled training data generated during an iteration is used to train the base caller during an immediate subsequent iteration.
A method of progressively training a base caller is disclosed. The method includes initially training a base caller, and generating labelled training data using the initially trained base caller; and (i) further training the base caller with analyte comprising organism base sequences, and generating labelled training data using the further trained base caller. The method includes iteratively further training the base caller by repeating step (i) for N iterations, which includes further training the base caller for N1 iterations of the N iterations with analyte comprising a first organism base sequence, and further training the base caller for N2 iterations of the N iterations with analyte comprising a second organism base sequence. A complexity of neural network configurations loaded in the base caller monotonically increases with the N iterations, and labelled training data generated during an iteration is used to train the base caller during an immediate subsequent iteration.
This disclosure describes methods, non-transitory computer readable media, and systems that can generate signal-to-noise-ratio metrics for clusters of oligonucleotides to which tagged nucleotide bases are added and utilize the signal-to-noise-ratio metrics to generate nucleotide-base calls and determine base-call quality. For example, the disclosed systems can generate the signal-to-noise-ratio metrics using scaling factors and noise levels associated with light signals detected from the clusters of oligonucleotides. The disclosed systems can utilize the signal-to-noise-ratio metrics to generate intensity-value boundaries for generating nucleotide-base-calls for the signals in accordance with one or more base-call-distribution models. Additionally, the disclosed systems can utilize a threshold to filter out signals detected from the clusters of oligonucleotides that have low signal-to-noise-ratio metrics. The disclosed systems can further utilize the signal-to-noise-ratio metrics to generate quality metrics for generated nucleotide-base calls
This disclosure describes methods, non-transitory computer readable media, and systems that can utilize a machine learning model to recalibrate nucleotide-base calls (e.g., variant calls) of a call-generation model. For instance, the disclosed systems can train and utilize a call-recalibration-machine-learning model to generate a set of predicted variant-call classifications based on sequencing metrics associated with a sample nucleotide sequence. Leveraging the set of variant-call classifications, the disclosed systems can further update or modify nucleotide-base calls (e.g., variant calls) corresponding to genomic coordinates. Indeed, the disclosed systems can generate an initial nucleotide-base call based on sequencing metrics for nucleotide reads of a sample sequence utilizing a call-generation model and further utilize a call-recalibration-machine-learning model to generate classification predictions for updating or recalibrating the initial nucleotide-base call from a subset of the same sequencing metrics or other sequencing metrics.
This disclosure describes methods, non-transitory computer readable media, and systems that can train a genome-location-classification model to classify or score genomic coordinates or regions by the degree to which nucleobases can be accurately identified at such genomic coordinates or regions. For instance, the disclosed systems can determine sequencing metrics for sample nucleic-acid sequences or contextual nucleic-acid subsequences surrounding particular nucleobase calls. By leveraging ground-truth classifications for genomic coordinates, the disclosed systems can train a genome-location-classification model to relate data from one or both of the sequencing metrics and contextual nucleic-acid subsequences to confidence classifications for such genomic coordinates or regions. After training, the disclosed systems can also apply the genome-location-classification model to sequencing metrics or contextual nucleic-acid subsequences to determine individual confidence classifications for individual genomic coordinates or regions and then generate at least one digital file comprising such confidence classifications for display on a computing device.
Some embodiments relate to methods and compositions for preparing combinatorially indexed beads. Some embodiments include sequential addition of different indexes to polynucleotides attached to beads. In some embodiments, indexes are added by chemical ligation, polymerase extension, ligation of partially double-stranded adaptors, or short splint ligation.
The present approach relates generally to image-based approaches for detecting deviations from a linear movement when scanning a surface. More particularly, the approach relates to the use of linear fiducials to detect, in real-time, deviations from a linear scan path during operation of a scanning imaging system. Such linear fiducials may include both sample sites and blank regions or sites or, in certain embodiments, may utilize elongated sample sites (e.g., linear features) within the linear fiducial.
Disclosed herein include systems, devices, and methods for determining a variable number tandem repeat (VNTR) status. Haplotypes of a VNTR can be determined using long sequence reads of reference samples aligned to the VNTR in a reference. Short reads of a test sample of a test subject can be aligned to the haplotypes determined using the long sequence reads to determine a VNTR status (e.g., one or more haplotypes or a genotype of the test subject) of the test subject based on the probability indications of the haplotypes.
G16H 50/20 - ICT specially adapted for medical diagnosis, medical simulation or medical data miningICT specially adapted for detecting, monitoring or modelling epidemics or pandemics for computer-aided diagnosis, e.g. based on medical expert systems
Disclosed herein include systems, devices, and methods for identifying recombinant variants (e.g., gene conversion variants) of genes such as GBA gene and CYP21A2 gene, the copy numbers of recombinant variants, and gene variant status (e.g., carrier, compound heterozygous, or homozygous).
Disclosed herein include systems, devices, and methods for determining repeat expansion status (e.g., pathogenic, carrier, and benign) of a locus of a gene of interest (e.g., at, or at about, chr4:39348424 of hg38 for RFC1). After aligning sequence reads to a sequence graph, the number of occurrences of repeat sequences satisfying predetermined criteria and the frequency of a pathogenic repeat sequence can be determined, which are in turn used to determine a repeat expansion status.
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.
An example of a flow cell includes a substrate; a plurality of reactive regions extending along the substrate; and a non-reactive region separating one of the plurality of reactive regions from an adjacent one of the plurality of reactive regions. Each of the plurality of reactive regions includes alternating first and second areas positioned along the reactive region. Each of the first areas includes a first primer set and each of the second areas includes a second primer set that is different than the first primer set. Either adjacent first and second areas directly abut each other, or) the first areas are positioned on protrusions and the second areas are positioned in depressions adjacent to the protrusions.
In an example of a method for making a flow cell, a metal material is sputtered over a transparent substrate including depressions separated by interstitial regions to form a metal film having a first thickness over the interstitial regions and having a second thickness over the depressions, the second thickness being about 30 nm or less and being at least 1/3 times smaller than the first thickness. A light sensitive material is deposited over the metal film; and the metal film is used to develop the light sensitive material through the transparent substrate to define an altered light sensitive material at a first predetermined region over the transparent substrate. The altered light sensitive material is utilized to generate a functionalized layer at the first predetermined region or at a second predetermined region over the transparent substrate.
B01J 19/00 - Chemical, physical or physico-chemical processes in generalTheir relevant apparatus
G03F 7/00 - Photomechanical, e.g. photolithographic, production of textured or patterned surfaces, e.g. printed surfacesMaterials therefor, e.g. comprising photoresistsApparatus specially adapted therefor
In an example of a method for making a flow cell, a light sensitive material is deposited over a resin layer including depressions separated by interstitial regions, wherein the depressions overlie a first resin portion having a first thickness and the interstitial regions overlie a second resin portion having a second thickness that is greater than the first thickness. A predetermined ultraviolet light dosage that is based on the first and second thicknesses is directed through the resin layer, whereby the light sensitive material overlying the depressions is exposed to ultraviolet light and the second resin portion absorbs the ultraviolet light, thereby defining an altered light sensitive material at a first predetermined region over the resin layer. The altered light sensitive material is utilized to generate a functionalized layer at the first predetermined region or at a second predetermined region over the resin layer.
B01J 19/00 - Chemical, physical or physico-chemical processes in generalTheir relevant apparatus
G03F 7/00 - Photomechanical, e.g. photolithographic, production of textured or patterned surfaces, e.g. printed surfacesMaterials therefor, e.g. comprising photoresistsApparatus specially adapted therefor
Nucleic acid techniques are disclosed. Embodiments include modified nucleotides 12 with oligonucleotide adapters 24 that are coupled via cleavable linkers 20. Incorporation of the modified nucleotide 12 at a 3' end of a nucleic acid permits end-adapterization via ligation of a free 5' end of the oligonucleotide adapter 24 to a 3' reactive group of the modified nucleotide 12 and cleavage at the cleavable linker 20 to liberate a free 3' end.
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.
C07H 19/20 - Purine radicals with the saccharide radical being esterified by phosphoric or polyphosphoric 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
C12P 19/34 - Polynucleotides, e.g. nucleic acids, oligoribonucleotides
73.
COMPOSITIONS AND METHODS FOR SEQUENCING BY SYNTHESIS
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.
Disclosed herein include systems, devices, and methods for grouping sequence reads and collapsing families of sequence reads that originate from the same DNA molecules using UMIs.
Nucleic acid amplification techniques are disclosed. Embodiments include generating concatenated nucleic acids using rolling circle amplification of templates, e.g., starting from a cDNA of a full-length mRNA or from synthetic templates, and sequencing and/or detecting the concatenated nucleic acids. In some embodiments, the technology disclosed includes amplification reactions that include CRISPR-Cas interactions that generate primers as a result of the CRISPR-Cas interactions, whereby primers are in turn used as part of detectable amplification reactions. The disclosed amplification techniques may use synthetic oligonucleotides or primers.
The technology disclosed relates to efficiently determining which atoms in a protein are nearest to voxels in a grid. The atoms have three-dimensional (3D) atom coordinates, and the voxels have 3D voxel coordinates. The technology disclosed generates an atom-to-voxels mapping that maps, to each of the atoms, a containing voxel selected based on matching 3D atom coordinates of a particular atom of the protein to the 3D voxel coordinates in the grid. The technology disclosed generates a voxel-to-atoms mapping that maps, to each of the voxels, a subset of the atoms. The subset of the atoms mapped to a particular voxel in the grid includes those atoms in the protein that are mapped to the particular voxel by the atom-to-voxels mapping. The technology disclosed includes using the voxel-to-atoms mapping to determine, for each of the voxels, a nearest atom in the protein.
G16B 15/00 - ICT specially adapted for analysing two-dimensional or three-dimensional molecular structures, e.g. structural or functional relations or structure alignment
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.
C12Q 1/6848 - Nucleic acid amplification reactions characterised by the means for preventing contamination or increasing the specificity or sensitivity of an amplification reaction
The technology disclosed relates to determining pathogenicity of variants. In particular, the technology disclosed relates to generating amino acid-wise distance channels for a plurality of amino acids in a protein. Each of the amino acid-wise distance channels has voxel-wise distance values for voxels in a plurality of voxels. A tensor includes the amino acid-wise distance channels and at least an alternative allele of the protein expressed by a variant. A deep convolutional neural network determines a pathogenicity of the variant based at least in part on processing the tensor. The technology disclosed further augments the tensor with supplemental information like a reference allele of the protein, evolutionary conservation data about the protein, annotation data about the protein, and structure confidence data about the protein.
G16B 15/00 - ICT specially adapted for analysing two-dimensional or three-dimensional molecular structures, e.g. structural or functional relations or structure alignment
G16B 20/00 - ICT specially adapted for functional genomics or proteomics, e.g. genotype-phenotype associations
A system includes at least a voxelizer, an alternative allele encoder, an evolutionary conservation encoder, and a convolutional neural network. The voxelizer accesses a three-dimensional structure of a reference amino acid sequence of a protein and fits a three-dimensional grid of voxels on atoms in the three-dimensional structure on an amino acid-basis to generate amino acid-wise distance channels. The alternative allele encoder encodes an alternative allele sequence to each voxel in the three-dimensional grid of voxels. The evolutionary conservation encoder encodes an evolutionary conservation sequence to each voxel in the three-dimensional grid of voxels. The convolutional neural network applies three-dimensional convolutions to a tensor that includes the amino acid-wise distance channels encoded with the alternative allele sequence and respective evolutionary conservation sequences and determines a pathogenicity of a variant nucleotide based at least in part on the tensor.
G16B 15/00 - ICT specially adapted for analysing two-dimensional or three-dimensional molecular structures, e.g. structural or functional relations or structure alignment
G16B 20/00 - ICT specially adapted for functional genomics or proteomics, e.g. genotype-phenotype associations
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.
The technology disclosed relates to systems and methods for diagnosing system malfunction and isolating a cause of system malfunction. The method includes applying preprocessors to time series data. The preprocessors detect time series discontinuities, drift, lack of expected correlation, and trends. The method includes feeding, for at least one image channel in one sequencing run, at least part of the output of the preprocessors to a trained tree-based classifier and receiving a classification of the particular sequencing run as abnormal. The abnormal classification can indicate a system malfunction. The method includes feeding at least part of the output of the preprocessors for the abnormal sequencing run to an expert rule system. The expert rule system can isolate a root cause of the system malfunction to a particular subcomponent in need of adjustment or replacement. The method can generate a notification of the particular subcomponent causing the system malfunction.
Described herein are standards and methods of normalizing amplicon size bias. These standards may comprise unique molecular identifiers. In some embodiments, the standards and methods are for use with next generation sequencing (NGS) assays. Also described herein are methods for quantifying DNA damage in a sample comprising DNA using fluorescence or for determining the presence of DNA damage in a library.
Described herein are compositions and methods for preparing double-stranded complementary DNA (cDNA) from RNA. In some embodiments, these methods allow isothermal preparation of cDNA. In some embodiments, these methods allow mesophilic or thermostable preparation of cDNA. Also described herein are compositions and methods for preparing cDNA and a library of double-stranded cDNA fragments in a single reaction vessel.
A neural network processes sequencing images on a patch-by-patch basis for base calling. The sequencing images depict intensity emissions of a set of analytes. The patches depict the intensity emissions for a subset of the analytes and have undiverse intensity patterns due to limited base diversity. The neural network has convolution filters that have receptive fields confined to the patches. The convolution filters detect intensity patterns in the patches with losses in detection due to the undiverse intensity patterns and confined receptive fields. An intensity contextualization unit determines intensity context data based on intensity values in the images. The data flow logic appends the intensity context data to the sequencing images to generate intensity contextualized images. The neural network applies the convolution filters on the intensity contextualized images and generates base call classifications. The intensity context data in the intensity contextualized images compensates for the losses in detection.
In one aspect, the disclosed technology relates to systems and methods for sequencing polynucleotides. In one embodiment, the disclosed technology relates to a nanopore sensor device for identifying nucleotides, the nanopore sensor device including: one or more cis wells; one or more cis electrodes associated with the one or more cis wells; a plurality of trans wells, each of the plurality of trans wells separated from the one or more cis wells by a lipid or solid-state membrane having a nanopore; a plurality of field effect transistors (FETs), each of the plurality of FETs associated with one of the plurality of trans wells; an electrical source configured to provide alternating current (AC) inputs between the one or more cis electrodes and the source terminals of the plurality of FETs; and a controller operably coupled to the plurality of FETs, the controller configured to measure AC responses of the plurality of FETs, wherein the AC responses depend on the identities of the nucleotides within or near the nanopores.
The disclosure relates to methods, compositions, and kits for the selective depletion of non-desirable fragments from amplified libraries using blocking oligonucleotides.
C12Q 1/6848 - Nucleic acid amplification reactions characterised by the means for preventing contamination or increasing the specificity or sensitivity of an amplification reaction
87.
MACHINE-LEARNING MODEL FOR DETECTING A BUBBLE WITHIN A NUCLEOTIDE-SAMPLE SLIDE FOR SEQUENCING
Methods, systems, and non-transitory computer readable media are disclosed for accurately and efficiently detect when bubbles impact nucleic-acid-sequencing runs based on data captured during (or derived from) base calls during sequencing runs. In particular, in one or more embodiments, the disclosed systems receive data identifying nucleobase calls and data identifying quality metrics for the nucleobase calls during sequencing cycles. Based on particular nucleobase calls and threshold markers for the quality metrics, the disclosed system utilizes a machine-learning-model to detect a presence of a bubble in a nucleotide-sample slide. Beyond simply detecting the presence of a bubble, the disclosed system can also classify different detected bubbles, such as air bubbles, oil bubbles, or ghost bubbles, or other outputs during sequencing. By utilizing call data and quality metrics, the disclose system can use readily available sequencing data in a platform-agnostic approach to detect bubbles using a uniquely trained machine-learning model.
Disclosed herein is a modified transposon end sequence comprising a mosaic end sequence, wherein the mosaic end sequence comprises one or more mutation as compared to a wild-type mosaic end sequence, wherein the mutation comprises a substitution with a uracil, an inosine, a ribose, an 8-oxoguanine, a thymine glycol, a modified purine, or a modified pyrimidine. -Also disclosed are transposome complexes comprising these modified transposon end sequences and methods of library preparation using these modified transposon end sequences.
Polypeptide nanopores synthetically functionalized with positively charged species, and methods of making and using the same, are provided herein. In some examples, a polypeptide nanopore includes a first side, a second side, a channel extending through the first and second sides, and a mutated amino acid residue. The mutated amino acid residue may be synthetically functionalized with a positively charged species that inhibits translocation of cations through the channel.
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.
Apparatus and methods for transmitting light are disclosed. In an implementation, an apparatus includes a collimator at an input end positioned to receive an input beam from a fiber beam source and to produce a collimated beam. The apparatus further includes a beam shaping group having one or more optical elements and positioned to receive the collimated beam from the collimator and format the collimated beam into a shaped propagation beam having a substantially rectangular cross-section in a far field. The apparatus further includes an objective stage for optically probing a sample, such as a flow cell, using substantially rectangular cross-section sampling beam, where fluorescence from the sample is captured by a line sensor for detecting properties of the sample, such as chemical reactions therein.
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.
A system for base calling includes memory storing a topology of a neural network, a plurality of weights sets, and sensor data for a series of sensing cycles. Sequencing events span temporal progression of the base calling operation through subseries of sensing cycles, and spatial progression of the base calling operation through locations on a biosensor. A configurable processor is configured to load the topology on the configurable processor, select a weight set in dependence upon a subject subseries of sensing cycles and/or a subject location on the biosensor, load subject sensor data for the subject subseries of sensing cycles and the subject location on the processing elements, configure the topology using the selected weight set, and cause the neural network to process the subject sensor data to produce base call classification data for the subject subseries and the subject location.
Analyzing expression of protein-coding variants in cells is provided herein. A method may include replacing a protein coding-region of the DNA in a cell with a donor vector including a variant of the protein-coding region and a first barcode identifying that variant. The cell may generate mRNA including an expression of the variant and an expression of the first barcode. A second barcode corresponding to the cell may be coupled to the mRNA. The mRNA, having the second barcode coupled thereto, may be reverse transcribed into complementary cDNA. The cDNA may be sequenced. The donor vector or cDNA may be sequenced using amplicon sequencing. The donor vector sequence and the cDNA sequence may be correlated to identify the variant and the cell's expression of the variant.
A structured illumination microscopy (SIM) system comprises: a light source; a light-structuring component to provide light from the light source with a SIM pattern for performing illuminations of a sample at a substrate having a substrate pattern, wherein a pitch of the SIM pattern is based on a characteristic of the substrate pattern; and an image sensor to detect emissions that the sample generates in response to the illuminations.
Genomic library preparation using Cas-gRNA RNPs, and targeted epigenetic assays, are provided herein. Some compositions include, from a first species, substantially only single- stranded polynucleotides; from a second species, substantially only double-stranded polynucleotides; and amplification primers ligated to ends of the second double-stranded polynucleotides and substantially not ligated to any ends of the first double-stranded polynucleotides. Some compositions include first and second molecules of a target polynucleotide having a sequence, the first molecule having a first end at a first subsequence, the second molecule having a first end at a second subsequence, wherein the first subsequence only partially overlaps with the second subsequence. Some examples provide a composition that includes a target polynucleotide and a first fusion protein including a Cas- gRNA RNP coupled to a transposase having an amplification adapter coupled thereto. The Cas-gRNA RNP may be hybridized to a subsequence in the target polynucleotide.
In some examples, a method of coupling oligonucleotides to a polymer is provided. Inactive moieties in a first region of a polymer may be selectively irradiated with light, while inactive moieties in a second region of the polymer are not irradiated, to generate first active moieties in the first region of the polymer. The first active moieties may be coupled to first oligonucleotides. The inactive moieties in the second region of the polymer may be irradiated with light to generate second active moieties in the second region of the polymer. The second active moieties may be coupled to second oligonucleotides.
There is set forth herein, in one example, an apparatus. The apparatus can comprise, for example: a first reaction site and a second reaction site over a single pixel. There is set forth herein, in one example, a method. The method can include, for example: detecting a signal emitted from a first reaction site and a second reaction site; determining the identity of a first analyte of interest in a first reaction site using an amplitude of the detected signal; and determining the identity of a second analyte of interest in a second reaction site using the amplitude of the detected signal.
An example of a resin composition includes an epoxy resin matrix and functionalized silica nanoparticles incorporated into the epoxy resin matrix. The functionalized silica nanoparticles have an average particle size up to about 10 nm. The resin composition is imprintable using nanoimprint lithography. The resin composition may be used to manufacture a patterned substrate for a flow cell.
C08L 83/06 - Polysiloxanes containing silicon bound to oxygen-containing groups
G03F 7/00 - Photomechanical, e.g. photolithographic, production of textured or patterned surfaces, e.g. printed surfacesMaterials therefor, e.g. comprising photoresistsApparatus specially adapted therefor
C12M 1/40 - Apparatus specially designed for the use of free, immobilised, or carrier-bound enzymes, e.g. apparatus containing a fluidised bed of immobilised enzymes
C12Q 1/68 - Measuring or testing processes involving enzymes, nucleic acids or microorganismsCompositions thereforProcesses of preparing such compositions involving nucleic acids
Provided herein include various examples of a flow cell and methods for forming aspects of flow cell. The method may include applying a first adhesive to a substrate. The method may include orienting a die on the first adhesive. The method may also include orienting a package on the first adhesive. The package includes a die and a top surface of the die comprises an active surface and electrical contact points. Surfaces adjacent to the active surface on at least two opposing sides of the active surface form fanout regions for utilization in a fluidic path of the flow cell. The method further may include applying a second adhesive to a part of the package and attaching a lid to the second adhesive to define a fluidic flow-cell cavity below the lid and above a surface comprising the active surface and the fanout regions.