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Небесная энциклопедия

Космические корабли и станции, автоматические КА и методы их проектирования, бортовые комплексы управления, системы и средства жизнеобеспечения, особенности технологии производства ракетно-космических систем

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Мониторинг СМИ

Мониторинг СМИ и социальных сетей. Сканирование интернета, новостных сайтов, специализированных контентных площадок на базе мессенджеров. Гибкие настройки фильтров и первоначальных источников.

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Применить Всего найдено 22475. Отображено 100.

12-01-2012 дата публикации

Methods for determining fraction of fetal nucleic acids in maternal samples

Номер: US20120010085A1

Автор: David A. COMSTOCK, Gabrielle HEILEK, Manjula CHINNAPPA, Michael Hunkapiller, Richard P. Rava, Yue-Jen Chuu

Принадлежит: Artemis Health Inc, Verinata Health Inc

The invention provides compositions and methods for determining the fraction of fetal nucleic acids in a maternal sample comprising a mixture of fetal and maternal nucleic acids. The fraction of fetal nucleic acids can be used in determining the presence or absence of fetal aneuploidy.

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Номер записи: 1

01-03-2012 дата публикации

OLIGONUCLEOTIDES FOR DETECTING E. coli O157:H7 STRAINS AND USE THEREOF

Номер: US20120052494A1

Автор: Jason Opdyke, Jun Li, Win Den Cheung

Принадлежит: Samsung Techwin Co Ltd

Oligonucleotides, a kit, and a method for detecting E. coli O157:H7 strains are provided. According to the kit for detecting E. coli O157:H7 strains and the method of detecting E. coli O157:H7 strains by using the kit, the results of the detection can be rapidly identified with a reduced number of copies of a sample in real-time.

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Номер записи: 2

08-03-2012 дата публикации

METHOD OF DIAGNOSING POOR SURVIVAL PROGNOSIS COLON CANCER USING miR-203

Номер: US20120058914A1

Автор: Aaron J. Schetter, Carlo M. Croce, Curtis C. Harris

Принадлежит: Ohio State University Research Foundation, US Department of Health and Human Services

The present invention provides novel methods and compositions for the diagnosis and treatment of colon cancers. In particular, the present invention provides diagnostics and prognostics for colon (including colon adenocarcinoma) cancer patients, wherein the methods related to measuring miR levels can predict poor survival. The invention also provides methods of identifying inhibitors of tumorigenesis.

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Номер записи: 3

15-03-2012 дата публикации

Modified nucleotide and real-time polymerase reaction using the same

Номер: US20120064531A1

Автор: Dae-Ro AHN

Принадлежит: Korea Advanced Institute of Science and Technology KAIST

The present invention relates to a modified nucleotide and real-time polymerase reaction using the nucleotide. Specifically, the present invention relates to a fluorescence material linked-nucleotide, a composition for real-time polymerase reaction comprising the nucleotide, an analysis kit and an analysis method. In the present invention, the fluorescence material linked-nucleotide serves the dual roles of producing fluorescence signal as well as being used as a substrate. Therefore, the present invention is economically advantageous because it is unnecessary to prepare probes, but can be applied to analyze various real-time polymerase reactions such as PCR, RCA and isothermal polymerization reaction, and shows higher quality of performance than the past methods.

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Номер записи: 4

29-03-2012 дата публикации

Method for Detecting and Quantitating Multiple-Subcellular Components

Номер: US20120075453A1

Автор: Michael Kilpatrick, Triantafyllos Tafas

Принадлежит: Ikonisys Inc

A method for detecting and quantitating multiple and unique fluorescent signals from a cell sample is provided. The method combines immunohistochemistry and a fluorescent-labeled in situ hybridization techniques. The method is useful for identifying specific subcellular components of cells such as chromosomes and proteins.

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Номер записи: 5

12-04-2012 дата публикации

Composition and methods for rapid detection of hiv by loop-mediated isothermal amplification (lamp)

Номер: US20120088244A1

Автор: Donna Rudolph, Kelly Curtis, S. Michele Owen

Принадлежит: US Department of Health and Human Services

Methods and compositions for detection of HIV nucleic acids in a sample, such as a biological sample obtained from a human subject, are provided according to embodiments of the present invention which include providing a reaction mixture including at least one LAMP, accelerated LAMP, RT-LAMP or RT-accelerated LAMP assay primer set specific for HIV-I or HIV-2 nucleic acids and the biological sample to be tested for presence of HIV-I and/or HIV-2 nucleic acids; incubating the reaction mixture under conditions suitable to produce a LAMP assay reaction product; and detecting the reaction product. Primers and primer sets for use in LAMP assays of HIV-I or HIV-2 nucleic acids are provided.

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Номер записи: 6

28-06-2012 дата публикации

Simultaneous determination of aneuploidy and fetal fraction

Номер: US20120165203A1

Автор: David A. COMSTOCK, Gabrielle HEILEK, Manjula CHINNAPPA, Richard P. Rava, Stephen Quake

Принадлежит: Verinata Health Inc

The invention provides compositions and methods for simultaneously determining the presence or absence of fetal aneuploidy and the relative amount of fetal nucleic acids in a sample obtained form a pregnant female. The method encompasses the use of sequencing technologies and exploits the occurrence of polymorphisms to provide a streamlined noninvasive process applicable to the practice of prenatal diagnostics.

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Номер записи: 7

30-08-2012 дата публикации

Arrays of microparticles and methods of preparation thereof

Номер: US20120220495A1

Автор: Chiu Wo Chau, Hui Huang, Jiacheng Yang, Michael Seul, Sukanta Banerjee, Ye Hong

Принадлежит: Bioarray Solutions Ltd

This invention provides high unit density arrays of microparticles and methods of assembling such arrays. The microparticles in the arrays may be functionalized with chemical or biological entities specific to a given target analyte. The high unit density arrays of this invention are formed on chips which may be combined to form multichip arrays according to the methods described herein. The chips and/or multichip arrays of this invention are useful for chemical and biological assays.

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Номер записи: 8

30-08-2012 дата публикации

Methods and Microfluidic Devices for the Manipulation of Droplets in High Fidelity Polynucleotide Assembly

Номер: US20120220497A1

Автор: Joseph Jacobson, Larry Li-Yang Chu, Senthil Ramu

Принадлежит: GEN 9 Inc

Methods and devices are provided for manipulating droplets on a support using surface tension properties, moving the droplets along a predetermined path and merging two droplets together enabling a number of chemical reactions. Disclosed are methods for controlling the droplets volumes. Disclosed are methods and devices for synthesizing at least one oligonucleotide having a predefined sequence. Disclosed are methods and devices for synthesizing and/or assembling at least one polynucleotide product having a predefined sequence from a plurality of different oligonucleotides having a predefined sequence. In exemplary embodiments, the methods involve synthesis and/or amplification of different oligonucleotides immobilized on a solid support, release of synthesized/amplified oligonucleotides in solution to form droplets, recognition and removal of error-containing oligonucleotides, moving or combining two droplets to allow hybridization and/or ligation between two different oligonucleotides, and further chain extension reaction following hybridization and/or ligation to hierarchically generate desired length of polynucleotide products.

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Номер записи: 9

27-09-2012 дата публикации

Method for detecting microorganisms belonging to mycoplasma pneumoniae and/or mycoplasma genitalium

Номер: US20120244544A1

Автор: Atsuko Minagawa, Hatsue Itagaki, Kazuyuki Sugiyama, Toyomasa Hiroshima, Yasushi Shimada, Yuki Mitobe

Принадлежит: LSI Medience Corp

A detection method and a detection kit for rapidly and specifically diagnosing Mycoplasma pneumoniae and/or Mycoplasma genitalium infections are provided. The DnaK of Mycoplasma pneumoniae or Mycoplasma genitalium is used as an indicator.

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Номер записи: 10

04-10-2012 дата публикации

Method for analyzing rna

Номер: US20120253687A1

Автор: Hitoshi Nobumasa, Osamu Nomura, Toshihiko Kuroda

Принадлежит: TORAY INDUSTRIES INC

A method of analyzing RNA extracted from a tissue or cell(s) fixed with a fixative, said method comprising a determining step of whether said RNA satisfies: B/A≦1 wherein A represents the weight ratio (%) of RNA within the range of 1000 to 4000 nucleotides with respect to the total weight of RNA as determined by electrophoresis; and B represents the weight ratio (%) of RNA within the range of more than 4000 nucleotides with respect to the total weight of RNA as determined by electrophoresis.

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Номер записи: 11

18-10-2012 дата публикации

Method for detecting a circularized dna, and use of said method for detecting mutations

Номер: US20120264630A1

Автор: Ismaïl Cisse, Ulrich Bockelmann, Virgile Viasnoff

Принадлежит: Individual

The present invention relates to a method for detecting a circularized single-stranded DNA by means of the isothermal hyperbranched rolling circle amplification technique, in which the primers used comprise a detectable barcode sequence and, optionally, a spacer which blocks polymerization by DNA polymerase. The present invention also relates to the use of said method for detecting a genetic polymorphism of one or more base pair(s).

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Номер записи: 12

01-11-2012 дата публикации

Kit for detection of microorganism

Номер: US20120277121A1

Автор: Shinichi Yoshida, Takashi Soejima

Принадлежит: Morinaga Milk Industry Co Ltd

A kit for use in a method for detecting live cells, injured cells and dead cells of a microorganism in a test sample by a nucleic acid amplification method is disclosed. The kit includes a cross-linker capable of cross-linking DNA by irradiation with light having a wavelength of 350 nm to 700 nm, medium, and a primer (s) for amplifying a DNA target region in the microorganism by a nucleic acid amplification method. The nucleic acid amplification method may be a PCR, LAMP, SDA, LCR or DNA microarray method. A cross-linker may be included in the kit such as ethidium monoazide, ethidium diazide, psoralen, 4,5′,8-trimethyl psoralen, or 8-methoxy psoralen.

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Номер записи: 13

22-11-2012 дата публикации

Determining codon distribution and/or base pair distance between codons in a nucleic acid

Номер: US20120295251A1

Автор: Adrian John Gibbs, Andrew Franklin, Christian Samundsett, Mark John Gibbs, Murali Nayudu, Sheba Khan, Terry John Murphy, Yafei Zhang

Принадлежит: EZYGENE Pty Ltd

The present invention relates to methods for the design and/or production of a probe or primer that is capable of hybridizing to a plurality of sites in a sample comprising nucleic acid. Furthermore, the present invention provides methods for detecting and amplifying nucleic acid using such a probe or primer, for example, for identification of a strain, species or genera. Probe or primer sequences are determined by reference to codon usage bias of a target nucleic acid. In addition, the present invention provides methods for determining codon distribution and/or base pair distance between codons in a nucleic acid.

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Номер записи: 14

20-12-2012 дата публикации

Materials and methods for detection of hpv nucleic acids

Номер: US20120322049A1

Автор: Anna K. Fulbright, Brian Lowe, Irina Nazarenko

Принадлежит: Qiagen Gaithersburg LLC

Provided are nucleic acids capable of hybridizing to HPV 16 and/or HPV 18 nucleic acids, in particular, mRNA encoding E2 and E6-7 gene products. Such nucleic acids are useful in methods of isolating RNA from a biological sample, methods and means for determining the presence of particular RNA splice-form variants in a biological sample, methods and means for determining the relative ratio of RNA ratios in a biological sample, methods and means for predicting the progression of precancerous cervical lesions, and methods and means for detecting disruption of genes or gene expression.

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Номер записи: 15

24-01-2013 дата публикации

Use of Ribozymes in the Detection of Adventitious Agents

Номер: US20130022964A1

Автор: Matthew C. Coffey

Принадлежит: Oncolytics Biotech Inc

The present invention provides a method of detecting adventitious agents in a composition comprising a microorganism by using ribozyme-expressing indicator cells, as well as indicator cells useful in such detection.

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Номер записи: 16

21-02-2013 дата публикации

Methods of Polynucleotide Detection

Номер: US20130045166A1

Автор: Allen T. Christian, G. David Grothaus, Jianping Xu, Larry A. Gilbertson, Mingqi Deng, R. Douglas Sammons, Ryan K. Bartlett, Sonya J. Franklin

Принадлежит: MONSANTO TECHNOLOGY LLC

The present invention provides methods of detecting for the presence of a polynucleotide in vivo. These methods are particularly useful for performing identification and/or analysis of samples or specimens in which it is impossible, impractical, or undesirable to move or remove them from their current environment. Methods of practicing the present invention for the purpose of identifying and/or analyzing transgenic plant tissue or cells, in addition to animal tissue or cells and bacterial cells are also provided.

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Номер записи: 17

28-03-2013 дата публикации

Probe, and polymorphism detection method using the same

Номер: US20130078631A1

Автор: Mariko Komori

Принадлежит: Arkray Inc

The present disclosure relates to a probe for detecting a polymorphism, a method of detecting a polymorphism, a method of evaluating the efficacy of a drug, and a reagent kit for detecting a polymorphism.

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Номер записи: 18

11-04-2013 дата публикации

Detection of Immune Cells, In Particular T Cells Through DNA-Methylation Analysis of the Genes CCR6 and BLR1

Номер: US20130089861A1

Автор: Sven Olek

Принадлежит: Individual

The present invention relates to a method, in particular an in vitro method, for identifying certain immune cells of a mammal, comprising analysing the methylation status of at least one CpG position in the gene CCR6 and/or BLR1 or an orthologous or paralogous gene thereof, and the use of DNA-methylation analysis of the genes of the proteins CCR6 and/or BLR1 for a detection and quality assurance and control of certain immune cells. In particular, the present invention relates to analysing the methylation status of at least one CpG position in the gene CCR6 in T cells. Furthermore, the present invention relates to a kit for performing the above methods, as well as to respective uses.

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Номер записи: 19

18-04-2013 дата публикации

Nmr systems and methods for the rapid detection of analytes

Номер: US20130095494A1

Автор: Lori Anne Neely

Принадлежит: T2 Biosystems Inc

This invention features systems and methods for the detection of analytes, and their use in the treatment and diagnosis of disease.

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Номер записи: 20

02-05-2013 дата публикации

Real-time redox sequencing

Номер: US20130109577A1

Автор: Jonas Korlach, Lei Sun, Stephen Turner

Принадлежит: Pacific Biosciences of California Inc

Real time redox sequencing methods, devices, and systems are described. Arrays of redox devices comprising one or two electrodes are used to provide sequence information about a template nucleic acid in a polymerase-template complex bound proximate to the electrode(s). A sequencing reaction mixture comprising nucleotide analogs comprising redox labels is introduced to the array of redox devices under conditions of polymerase mediated nucleic acid synthesis. The time sequence of incorporation of nucleotide analogs is determined by electrochemically identifying the redox labels of the nucleotide analogs that are incorporated into the growing strand.

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Номер записи: 21

16-05-2013 дата публикации

Method for producing rna-containing probe for detecting a target nucleotide

Номер: US20130122503A1

Автор: Hiroyuki Mukai, Junko Yamamoto, Kiyozo Asada, Tomoo Inden, Tooru Suzuki, Toshiharu Ohba

Принадлежит: Takara Bio Inc

An object of the present invention is to provide a simple and useful method for producing an RNA-containing probe for detecting a target nucleotide, a simple and useful method, device, and system for processing nucleotide sequence information, and a simple and useful method for detecting a target nucleotide. The present invention provides a method for processing nucleotide sequence information, the method comprising the step of generating partial nucleotide sequences which has 7 to 14 nucleotides and a Tm value of 25 to 40° C. and in which a target nucleotide or a nucleotide adjacent to the target nucleotide is located at a position between 3 and 5 nucleotides from the 3′ or 5′ end. The method according to the present invention is useful for simply and efficiently producing an RNA-containing probe for detecting a target nucleic acid, without the basis of researchers' experiences or guess, and are extremely useful not only in the field of genetic engineering, but also in the field of medical research.

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Номер записи: 22

16-05-2013 дата публикации

Methods and devices for manipulation of target cells using a combined application of acoustical and optical radiations

Номер: US20130122564A1

Автор: Boris Krasovitski, Eitan Kimmel, Shy Shoham

Принадлежит: Technion Research and Development Foundation Ltd

A method of applying a nondestructive mechanical force on one or more cells in aqueous environment by inducing heat generated acoustic pressure pulses. The method comprises providing an energy transmission pattern to induce the applying of a desired nondestructive mechanical force on a at least one cell by forming a plurality of heat generated acoustic pressure pulses in an aqueous environment and instructing the radiation of a target area in proximity to the at least one cell in the aqueous environment with light energy according to the energy transmission pattern.

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Номер записи: 23

30-05-2013 дата публикации

Rapid nucleic acid purification

Номер: US20130137107A1

Автор: Minsu KO, Siseok Lee, Young Cheol Kim

Принадлежит: Nanohelix Co Ltd

Provided is a method for rapid nucleic acid purification, and the method for rapid nucleic acid isolation according to the present invention is very useful in diagnosing causes of disease or detecting a target gene; can be used in molecular diagnosis of causes of disease more rapidly and conveniently, as compared with the existing nucleic acid isolation method requiring complicated and special equipment; does not require skills therefor, thereby allowing an ordinary person to personally conduct isolation of nucleic acid for analyzing causes of disease and further solving the existing inconvenience caused by directly going to the hospitals or health clinical centers; and can analyze causes of disease more promptly.

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Номер записи: 24

06-06-2013 дата публикации

Optical instrument comprising multi-notch beam splitter

Номер: US20130143310A1

Автор: Eugene F. Young, Mark F. Oldham

Принадлежит: APPLIED BIOSYSTEMS LLC

An instrument is provided that can monitor nucleic acid sequence amplification reactions, for example, PCR amplification of DNA and DNA fragments. The instrument includes a multi-notch filter disposed along one or both of an excitation beam path and an emission beam path. Methods are also provided for monitoring nucleic acid sequence amplifications using an instrument that includes a multi-notch filter disposed along a beam path.

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Номер записи: 25

04-07-2013 дата публикации

Method of dna sequencing by polymerisation

Номер: US20130171636A1

Автор: David Bensimon, Fang-Yuan Ding, Jean-Francois Allemand, Maria Manosas, Vincent Croquette

Принадлежит: ECOLE NORMALE SUPERIEURE

Described herein is a method for determining a nucleic acid sequence, said method comprising: a) denaturing a double-stranded nucleic acid molecule corresponding to the said nucleic acid sequence; b) hybridizing a single-stranded nucleic acid molecule, the primer, with the said denatured double-stranded nucleic acid molecule; c) applying a tension to the hybridized primer/double-stranded nucleic acid molecule obtained in b); d) incubating the hybridized primer/double-stranded nucleic acid molecule obtained in b) with a polymerase in conditions which will lead to at least one pause in replication; and e) determining a position of the said pause in replication with respect to one end of the double-stranded nucleic acid.

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Номер записи: 26

18-07-2013 дата публикации

Microfluidic devices

Номер: US20130183659A1

Автор: Darren R. Link, David Marran, Jonathan M. Rothberg, Michael Weiner

Принадлежит: Raindance Technologies Inc

The present invention provides novel microfluidic substrates and methods that are useful for performing biological, chemical and diagnostic assays. The substrates can include a plurality of electrically addressable, channel bearing fluidic modules integrally arranged such that a continuous channel is provided for flow of immiscible fluids.

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Номер записи: 27

05-09-2013 дата публикации

Capture of target dna and rna by probes comprising intercalator molecules

Номер: US20130230856A1

Автор: Jan Gorm Lisby, Nina JØHNK, Uffe Vest Schneider

Принадлежит: Quantibact AS

The present invention relates to a technology for specific capture of single stranded target Polynucleotide by a complementary probe comprising one or more intercalator molecules. The method further involves removal of one or more types of bases in the single stranded target Polynucleotide prior to interaction with the complementary probe. This results in generation of one or more abasic sites which can interact with and/or into where the intercalator molecule can be inserted.

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Номер записи: 28

12-09-2013 дата публикации

Separation of pyrophosphate release and pyrophosphate detection

Номер: US20130237434A1

Автор: Bernard Hirschbein, Filiz Gorpe-Yasar

Принадлежит: Illumina Inc

The present technology relates to methods and systems for detection of pyrophosphate. As such, disclosed herein are methods and systems that permit improved pyrophosphate detection. Also disclosed herein are methods and systems which utilize improved pyrophosphate detection for nucleotide sequencing.

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Номер записи: 29

19-09-2013 дата публикации

Compositions and methods for sequencing nucleic acids

Номер: US20130244886A1

Автор: Jason Richard Betley, Jonathan Mark Boutell, Niall Anthony Gormley, Roberto Rigatti

Принадлежит: Illumina Inc

Disclosed herein are compositions and methods for sequencing nucleic acids.

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Номер записи: 30

17-10-2013 дата публикации

Nucleic acid sample preparation

Номер: US20130273640A1

Автор: David CHARLOT, David Liu, Eugene Tu, Irina DOBROVOLSKAYA, James MCCANNA, Kai Yang, Lucas Kumosa, Paul Swanson, Rajaram Krishnan, Robert Turner

Принадлежит: Biological Dynamics Inc

The present invention includes methods, devices and systems for isolating a nucleic acid from a fluid comprising cells. In various aspects, the methods, devices and systems may allow for a rapid procedure that requires a minimal amount of material and/or results in high purity nucleic acid isolated from complex fluids such as blood or environmental samples.

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Номер записи: 31

24-10-2013 дата публикации

Methods and apparatus for particle introduction and recovery

Номер: US20130277223A1

Автор: Andrea Marziali, David John Broemeling, Dylan Corey Gunn, Joel Pel, Peter Jason Eugster

Принадлежит: University of British Columbia

Particles may be injected into a matrix for concentration by scodaphoresis using a quadrupole injection field. Particles may be injected from two or more sample chambers simultaneously. Particle injection may be performed simultaneously with performing scodaphoresis. In some embodiments the particles are concentrated into a well containing fluid. The well can extend out of a plane of the matrix. Altering the relative phases of components of a scodaphoresis field permits concentration of selected particles and exclusion of other particles. Scodaphoresis methods may be applied to DNA, other bio-molecules and other particles.

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Номер записи: 32

31-10-2013 дата публикации

Nanoscale calorimeter on chip and related methods and devices

Номер: US20130288386A1

Автор: Chung Wah Fon, Michael L. Roukes

Принадлежит: California Institute of Technology CalTech

An article comprising: an array of calorimeter devices, wherein the device comprises: at least one fluidic enclosure disposed on a microfluidic chip, wherein the fluidic enclosure is substantially gas impermeable; at least one first chamber and at least one second chamber, wherein the first chamber and the second chamber are disposed within and enclosed by the fluidic enclosure, wherein the first chamber and the second chamber are not vacuum encapsulated; at least two microfluidic channels connected to the first chamber and at least two microfluidic channels connected to the second chamber; and at least one thermal sensor disposed between the chip and the first and second chambers, wherein the thermal sensor is adapted to measure a temperature differential between the first and second chambers. Examples include DSC and TSA devices. Biological binding and melting experiments can be done with high sensitivity.

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Номер записи: 33

12-12-2013 дата публикации

Method for predicting the outcome of colon cancer by analysing mirna expression

Номер: US20130331291A1

Автор: Bernhard Mlecnik, Franck Pages, Herve Fridman, Jérôme GALON

Принадлежит: Assistance Publique Hopitaux de Paris APHP, Universite Paris Descartes Paris 5

The present invention relates to a method for predicting the outcome of a cancer. More particularly, the present invention relates to a method for predicting the outcome of a cancer in a patient comprising a step consisting of determining the expression level of a miRNA cluster in a sample obtained from said patient, wherein said miRNA cluster comprises: -miR.609 or, -miR.518c or, -miR.520f or, -miR.220a or, -miR.362 or, -miR.29a or, -miR.660 or, -miR.603 or, -miR.558 or, -miR519b or, -miR.494 or, -miR.130a or, -miR.639.

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Номер записи: 34

26-12-2013 дата публикации

Nucleic Acid Analysis by Random Mixtures of Non-Overlapping Fragments

Номер: US20130345070A1

Автор: Radoje Drmanac

Принадлежит: Callida Genomics Inc

The invention provides methods and kits for ordering sequence information derived from one or more target polynucleotides. In one aspect, one or more tiers or levels of fragmentation and aliquoting are generated, after which sequence information is obtained from fragments in a final level or tier. Each fragment in such final tier is from a particular aliquot, which, in turn, is from a particular aliquot of a prior tier, and so on. For every fragment of an aliquot in the final tier, the aliquots from which it was derived at every prior tier is known, or can be discerned. Thus, identical sequences from overlapping fragments from different aliquots can be distinguished and grouped as being derived from the same or different fragments from prior tiers. When the fragments in the final tier are sequenced, overlapping sequence regions of fragments in different aliquots are used to register the fragments so that non-overlapping regions are ordered. In one aspect, this process is carried out in a hierarchical fashion until the one or more target polynucleotides are characterized, e.g. by their nucleic acid sequences, or by an ordering of sequence segments, or by an ordering of single nucleotide polymorphisms (SNPs), or the like.

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Номер записи: 35

09-01-2014 дата публикации

Nucleic Acid Analysis by Random Mixtures of Non-Overlapping Fragments

Номер: US20140011688A1

Автор: Radoje Drmanac

Принадлежит: Callida Genomics Inc

The invention provides methods and kits for ordering sequence information derived from one or more target polynucleotides. In one aspect, one or more tiers or levels of fragmentation and aliquoting are generated, after which sequence information is obtained from fragments in a final level or tier. Each fragment in such final tier is from a particular aliquot, which, in turn, is from a particular aliquot of a prior tier, and so on. For every fragment of an aliquot in the final tier, the aliquots from which it was derived at every prior tier is known, or can be discerned. Thus, identical sequences from overlapping fragments from different aliquots can be distinguished and grouped as being derived from the same or different fragments from prior tiers. When the fragments in the final tier are sequenced, overlapping sequence regions of fragments in different aliquots are used to register the fragments so that non-overlapping regions are ordered. In one aspect, this process is carried out in a hierarchical fashion until the one or more target polynucleotides are characterized, e.g. by their nucleic acid sequences, or by an ordering of sequence segments, or by an ordering of single nucleotide polymorphisms (SNPs), or the like.

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Номер записи: 36

30-01-2014 дата публикации

Biomarkers for predicting sensitivity to cancer treatments

Номер: US20140031259A1

Автор: Elizabeth Punnoose, Kui Lin, Somasekar Seshagiri

Принадлежит: Genentech Inc

Methods for predicting sensitivity to cancer treatments by assessing biomarkers and combinations of biomarkers include evaluating the presence of mutations to Akt, wherein the presence of said mutations correlates with the sensitivity of Akt inhibitors.

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Номер записи: 37

27-02-2014 дата публикации

Binary DNA Probe for Fluorescent Analysis of Nucleic Acids

Номер: US20140057252A1

Автор: Dmitry Kolpashchikov

Принадлежит: Columbia University of New York

The invention is directed to binary oligonucleotide probes for nucleic analysis, which probes can be made of DNA or RNA that recognize nucleic acid analytes (both DNA and RNA) with unprecedented high selectivity under mild conditions and are highly sensitive to single nucleotide mismatches (SNP single nucleotide polymorphisms) without PCR amplification. In one group, the binary probes indicate that they have hybridized to a particular nucleic analyte by binding to a molecular beacon that gives off a fluorescent signal. A second group of binary probes bind to a dye such as malachite green, where upon hybridization to analyte the fluorescence of the dye increases dramatically and is easily detected and measured. The new binary probes require only about five minutes at room temperature to generate a detectable signal.

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Номер записи: 38

06-01-2022 дата публикации

CARBORHODAMINE COMPOUNDS AND METHODS OF PREPARATION THEREOF

Номер: US20220002549A1

Автор: Lukhtanov Eugeny A.

Принадлежит: ELITechGroup, Inc.

The carborhodamine dyes disclosed herein are novel reagents suitable for automated incorporation of carborhodamine dyes into oligonucleotides that can be used in detection methods for nucleic acid targets. This disclosure provides an efficient and simple process for the preparation of carborhodamine compounds and introduces previously unknown reagents for the automated synthesis of oligonucleotide-carborhodamine conjugates. 2. The method of claim 1 , wherein the solvent is 1 claim 1 ,2-dichloroethane and the Lewis acid is aluminum chloride.3. The method of claim 1 , wherein one or more hydrocarbon hydrogens of Ror Ris replaced with a linking group.4. The method of claim 1 , wherein R claim 1 , R claim 1 , R claim 1 , R claim 1 , R claim 1 , R claim 1 , R claim 1 , and Rform one or more 4- to 7-member ring systems by bridging between one or more of Rwith R claim 1 , Rwith R claim 1 , Rwith R claim 1 , Rwith R claim 1 , Rwith R claim 1 , and Rwith R.5. The method of claim 1 , wherein R claim 1 , R claim 1 , R claim 1 , R claim 1 , R claim 1 , R claim 1 , R claim 1 , and Rform one or more 4- to 7-member ring systems linked to one or more further 4- to 7-member ring systems claim 1 , and wherein the one or more further 4- to 7-member ring systems comprise up to 3 double bonds.6. The method of claim 5 , wherein the one or more further 4- to 7-member ring systems comprise heteroatoms.7. The method of claim 1 , wherein R claim 1 , R claim 1 , Rand Rform one or more 4- to 7-member ring systems by bridging between one or more of Rwith R claim 1 , Rwith R claim 1 , and Rwith R.8. The method of claim 1 , wherein R claim 1 , R claim 1 , Rand Rform one or more 4- to 7-member ring systems linked to one or more further 4- to 7-member ring systems claim 1 , and wherein the one or more further 4- to 7-member ring systems comprise up to 3 double bonds.9. The method of claim 8 , wherein the one or more further 4- to 7-member ring systems comprise heteroatoms.10. The method of claim 1 ...

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Номер записи: 39

06-01-2022 дата публикации

COMPETITIVE PROBES FOR ENGINEERING SIGNAL GENERATION

Номер: US20220002788A1

Автор: GOSCH Gregory, JACKY Lucien A.E., MENGE Karen L., RAJAGOPAL Aditya

Принадлежит:

A method of identifying a first target nucleic acid comprising, providing a sample comprising the first target nucleic acid, providing a first set of paired oligonucleotides with complementarity to the first target nucleic acid, the first set of paired oligonucleotides comprising a first ratio of (a) first competitive oligonucleotides to (b) first signal oligonucleotides comprising a signal tag, wherein the competitive oligonucleotides compete with the signal oligonucleotides for binding to the first target nucleic acid, amplifying the first target nucleic acid with the polymerase chain reaction, thereby degrading the first signal oligonucleotide and permitting generation of a first signal, generating the first signal, measuring intensity of the first signal, and correlating the intensity of the first signal to the first ratio, thereby identifying the first target nucleic acid. 1. Method of identifying a first target nucleic acid comprising ,providing a sample comprising the first target nucleic acid,providing a first set of paired oligonucleotides with complementarity to the first target nucleic acid, said first set of paired oligonucleotides comprising a first ratio of (a) first competitive oligonucleotides to (b) first signal oligonucleotides comprising a first signal tag,wherein the first competitive oligonucleotides compete with the first signal oligonucleotides for binding to the first target nucleic acid,amplifying the first target nucleic acid with the polymerase chain reaction, thereby degrading the first signal oligonucleotide and permitting generation of a first signal,generating the first signal,measuring intensity of the first signal, andcorrelating the intensity of the first signal to the first ratio, thereby identifying the first target nucleic acid.2. The method of claim 1 , wherein the sample comprises a second target nucleic acid.3. The method of claim 2 , further comprising identifying the second target nucleic acid by:providing a second set of ...

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Номер записи: 40

06-01-2022 дата публикации

TAGMENTATION-ASSOCIATED MULTIPLEX PCR ENRICHMENT SEQUENCING

Номер: US20220002793A1

Автор: AMBUR Ole Herman, ELLONEN Pekka, KRAUS CHRISTIANSEN Irene, LAGSTRÖM Sonja, LEPISTÖ Maja, MEISAL Roger, ROUNGE Trine

Принадлежит:

The present invention is related to methods for parallel sequencings of nucleic acid target sequences of interest, and in particular to massively parallel sequencing of nucleic acid sequences such as viral sequences that may have been integrated into a genome. For example, the methods, systems and kits provided herein may be used to enrich and sequence viral DNA sequences such as HPV and HIV sequences. 1. A method of amplifying a target nucleic acid sequence for use in a parallel sequencing method comprising:tagmenting a target nucleic sample to provide a plurality of tagmented sequences comprising a transposon adapter sequence at the ends of the tagmented sequences;contacting a first sample of the tagmented sequences with 1) a tag primer comprising a tag sequence portion that anneals to the transposon adapter sequence and a tag primer adapter portion, 2) a plurality of forward primers, each forward primer comprising a target sequence portion that anneals to a preselected portion of the sense strand of the target nucleic acid sequence and a forward primer sequencing portion, and 3) a sequencing primer comprising a portion that anneals to the forward sequencing portion of the forward primer and a sequencing primer adapter portion;performing a forward amplification reaction on the first sample of the tagmented sequences to provide a first library of amplicons spanning the target nucleic acid sequence;contacting a second sample of the tagmented sequences with 1) a tag primer comprising a tag sequence portion that anneals to the transposon sequence and a tag primer adapter portion, 2) a plurality of reverse primers, each reverse primer comprising a target sequence portion that anneals to a preselected portion of the antisense strand of the target nucleic acid sequence and a reverse primer sequencing portion, and 3) a sequencing primer comprising a portion that anneals to the reverse sequencing portion of the forward primer and a sequencing primer adapter portion; ...

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Номер записи: 41

07-01-2016 дата публикации

Methods and compositions for tagging and analyzing samples

Номер: US20160001248A1

Автор: Edward A. Hutchins, Hei-Mun Christina Fan

Принадлежит: Lineage Biosciences Inc USA

The invention relates to methods of tagging analytes in a sample.

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Номер записи: 42

02-01-2020 дата публикации

Droplet digital pcr chip

Номер: US20200001301A1

Автор: Jian Shi, Shuanghong Xiang, Xiaohui Song

Принадлежит: Pilot Gene Technologies (hangzhou) Co Ltd

The present invention discloses a droplet digital PCR chip. The droplet digital PCR chip includes at least one chip unit, each chip unit includes a chip body formed by bonding a top piece and a bottom piece, the chip body is internally provided with an inlet chamber, a droplet storage chamber, and an injection hole. The injection hole connects with the inlet chamber, a plurality of droplet generating channels are disposed between the inlet chamber and the droplet storage chamber, a height of the droplet generating channel is smaller than a height of the droplet storage chamber, an injection fluid is injected into the inlet chamber through the injection hole, and the injection fluid is emulsified and enters the droplet storage chamber at a junction of the droplet generating channels and the droplet storage chamber.

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Номер записи: 43

04-01-2018 дата публикации

METHODS OF CONSTRUCTING A CIRCULAR TEMPLATE AND DETECTING DNA MOLECULES

Номер: US20180002731A1

Автор: Tien Meng-Tsung, WU Mengchu

Принадлежит: Personal Genomics, Inc.

A method of constructing a circular template includes preparing a partially double stranded linear DNA molecule, and incubating the partially double stranded linear DNA molecule with a ligase capable of intra-molecular ligation of single stranded DNA molecules to generate a partially double stranded circular DNA molecule. A method of detecting DNA molecules includes the following steps. Target DNA molecules are isolated with probes to form partially double stranded linear DNA molecules. The partially double stranded linear DNA molecules are incubated with ligases capable of intra-molecular ligation of single stranded DNA molecules to generate partially double stranded circular DNA molecules. A circular sequencing of the partially double stranded circular DNA molecules is conducted by using the probes as primers. 1. A method of constructing a circular template comprising:preparing a partially double stranded linear DNA molecule, wherein the partially double stranded linear DNA molecule comprises single stranded protruding portions at both ends on a same strand; andincubating the partially double stranded linear DNA molecule with a ligase capable of intra-molecular ligation of single stranded DNA molecules to generate a partially double stranded circular DNA molecule.2. The method as claimed in claim 1 , wherein the step of preparing the partially double stranded linear DNA molecule comprises:providing a single stranded linear DNA molecule; andhybridizing a probe to the single stranded linear DNA molecule.3. The method as claimed in claim 2 , wherein the single stranded linear DNA molecule comprises 40 nucleotides to 500 nucleotides in length.4. The method as claimed in claim 2 , wherein the probe comprises 20 nucleotides to 120 nucleotides in length.5. The method as claimed in claim 2 , wherein the probe comprises a biotinylated probe.6. The method as claimed in claim 5 , after hybridizing the probe to the single stranded linear DNA molecule claim 5 , further ...

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Номер записи: 44

04-01-2018 дата публикации

METHODS FOR MOLECULAR DETECTION

Номер: US20180002734A1

Автор: Jackson George, Quan Christopher

Принадлежит:

This invention relates to methods for molecular detection, particularly to methods utilizing target-specific molecular probes. In exemplary embodiments, target-specific molecular probes include single-stranded deoxyribonucleic acid (DNA) or ribonucleic acid (RNA) aptamers. In general, the molecular probe may bind with relatively high specificity to a given target. In one aspect, a method for molecular detection comprises a molecular probe paired to a reporter molecule wherein the molecular probe impairs the amplification of the reporter molecule in the absence of the target molecule. 2. The method of claim 1 , wherein said at least one molecular probe is added prior to the addition of either of said at least one reporter molecule claim 1 , primer or both.3. The method of claim 2 , wherein said at least one molecular probe claim 2 , at least one reporter molecule and at least one primer are added simultaneously.4. The method of claim 1 , wherein said primer extension reaction comprises rolling circle amplification.5. The method of claim 1 , wherein said at least one molecule probe comprises a 3′ modification which is non-extensible by said primer extension reaction.6. The method of claim 1 , wherein said at least one primer shares at least a portion of its sequence in common with said at least one molecule probe.7. The method of claim 4 , wherein said rolling circle amplification is performed with a polymerase enzyme having strand displacement activity.8. The method of claim 1 , wherein said single-stranded nucleic acid product comprises a concatenation of the complement of said reporter molecule.9. The method of claim 1 , further comprising performing a quantification of said single-stranded nucleic acid product claim 1 , wherein said quantification correlates to the amount of said target present in said sample.10. The method of claim 9 , wherein said quantification comprises adding a dye that binds to said single-stranded nucleic acid product.11. The method of ...

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Номер записи: 45

04-01-2018 дата публикации

METHOD FOR IDENTIFYING THE SOURCE OF AN AMPLICON

Номер: US20180002751A1

Автор: Jesse Taco Peter, Van Eijk Michael Josephus Theresia, Van Tunen Adrianus Johannes

Принадлежит: Keygene N.V.

The present invention relates to a method for identifying the source of an amplicon, comprising: providing a plurality of pools of amplicons from different sources, wherein the amplicons from different sources are present in more than one pool, and wherein the amplicons in each pool are tagged with a unique pool-specific identifier; sequencing at least part of the amplicons that comprise the pool-specific identifiers; and assigning one or more of the amplicons to corresponding pools and/or sources using the pool-specific identifiers. 1. A method for identifying the source of an amplicon , comprising:(a) providing a plurality of pools of amplicons from different sources, wherein the amplicons from different sources are present in more than one pool, and wherein the amplicons in each pool are tagged with a unique pool-specific identifier;(b) sequencing at least part of the amplicons that comprise the pool-specific identifiers;(c) assigning one or more of the amplicons to corresponding pools and/or sources using the pool-specific identifiers.2. The method according to claim 1 , wherein the sequencing is carried out by high-throughput sequencing.3. The method according to claim 2 , wherein the high-throughput sequencing is performed on a solid support.4. The method according to claim 2 , wherein the high-throughput sequencing is based on Sequencing-by-Synthesis.5. The method according to claim 2 , wherein the high-throughput sequencing comprises the steps of:annealing the tagged nucleic acid fragments to beads, each bead annealing with a single tagged nucleic acid fragment;emulsifying the beads in water-in-oil micro reactors, each water-in-oil micro reactor comprising a single bead;performing emulsion PCR to amplify tagged nucleic acid fragments on the surface of beads,optionally, selecting and enriching beads comprising amplified tagged nucleic acid fragments;loading the beads in wells, each well comprising a single bead; andgenerating a pyrophosphate signal.6. The ...

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Номер записи: 46

02-01-2020 дата публикации

POLYNUCLEOTIDES FOR THE AMPLIFICATION AND DETECTION OF CHLAMYDIA TRACHOMATIS

Номер: US20200002752A1

Автор: DeDent Andrea, LEE Matt, MA Shuyuan, MAAMAR Hédia

Принадлежит:

The invention provides methods and compositions for the detection of in a test sample. Its presence or absence in the sample is determined by nucleic acid based testing methods using primers and/or probes and or molecular beacons that bind to the 23S ribosomal genes or gene transcripts. 1. A composition comprising a set of polynucleotides selected from the group consisting of Set-20 , Set-47 , and Set-74.2. The composition of claim 1 , further comprising a probe.3. The composition of claim 2 , wherein the probe is a labeled polynucleotide.4. The composition of claim 3 , wherein the labeled polynucleotide comprises a sequence selected from the group consisting of nucleotides 5-34 of SEQ ID NO: 97 claim 3 , nucleotides 5-31 of SEQ ID NO: 98 claim 3 , nucleotides 3-31 of SEQ ID NO: 99 claim 3 , nucleotides 5-30 of SEQ ID NO: 100 claim 3 , nucleotides 6-29 of SEQ ID NO: 101 claim 3 , nucleotides 5-26 of SEQ ID NO: 103 claim 3 , nucleotides 4-27 of SEQ ID NO: 104 claim 3 , nucleotides 7-30 of SEQ ID NO: 105 claim 3 , nucleotides 5-27 of SEQ ID NO: 106 claim 3 , nucleotides 6-29 of SEQ ID NO: 107 claim 3 , nucleotides 6-29 of SEQ ID NO: 108 claim 3 , nucleotides 6-29 of SEQ ID NO: 109 claim 3 , nucleotides 6-30 of SEQ ID NO: 110 claim 3 , nucleotides 8-31 of SEQ ID NO: 111 claim 3 , nucleotides 6-29 of SEQ ID NO: 112 claim 3 , nucleotides 8-31 of SEQ ID NO: 113 claim 3 , nucleotides 5-29 of SEQ ID NO: 114 claim 3 , nucleotides 5-29 of SEQ ID NO: 115 claim 3 , nucleotides 4-28 of SEQ ID NO: 116 claim 3 , nucleotides 5-30 of SEQ ID NO: 117 claim 3 , nucleotides 8-28 of SEQ ID NO: 118 claim 3 , nucleotides 8-30 of SEQ ID NO: 119 claim 3 , nucleotides 8-29 of SEQ ID NO: 120 claim 3 , nucleotides 4-33 of SEQ ID NO: 121 claim 3 , nucleotides 7-34 of SEQ ID NO: 122 claim 3 , nucleotides 9-34 of SEQ ID NO: 123 claim 3 , nucleotides 8-34 of SEQ ID NO: 124 claim 3 , nucleotides 6-28 of SEQ ID NO: 125 claim 3 , nucleotides 7-28 of SEQ ID NO: 126 claim 3 , nucleotides 3-27 of SEQ ID ...

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Номер записи: 47

02-01-2020 дата публикации

NUCLEIC ACID DETECTION METHOD

Номер: US20200002756A1

Автор: Egan Christopher, LAMBLE Henry John, LLOYD David

Принадлежит: Sense Biodetection Limited

The present invention relates to methods for the detection of nucleic acids of defined sequence and kits for use in said methods. The methods employ nicking agent(s), polymerase and oligonucleotide probes to produce probe fragments in the presence of a target nucleic acid. 2. A method according to wherein the second oligonucleotide probe (P2) is attached to a solid material or to a moiety that permits its attachment to a solid material.3. (canceled)4. A method according to wherein the probe (P1) derived strand of the double-stranded nucleic acid amplifier comprises one or more modifications that render it resistant to cleavage by the first and/or second nicking agent(s) claim 1 , such as a phosphorothioate linkage.5. (canceled)6. A method according to wherein the double-stranded nucleic acid amplifier contains two or more nicking agent cleavage sites.7. A method according to wherein the sample is also contacted with a target nucleic acid primer which optionally contains the cleavage site for the first nicking agent.8. A method according to wherein the first oligonucleotide probe (P1) comprises a complementarity region at its 3′ end that is capable of sequence specific hybridisation to the target nucleic acid; whereby claim 1 , following hybridisation of probe (P1) to the target nucleic acid claim 1 , extension of probe (P1) by the polymerase forms a double-stranded nucleic acid that contains the cleavage site for the first nicking agent within the target nucleic acid strand; and whereby the first nicking agent specifically recognises said double-stranded nucleic acid and cleaves said target nucleic acid strand to produce a primer that remains hybridised to the probe (P1) derived strand; and the polymerase extends the primer to produce the double-stranded nucleic acid amplifier which is cleaved in step a) (A).9. A method according to wherein probe (P1) contains the recognition sequence for a probe (P1) nicking agent and whereby the double-stranded nucleic acid ...

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Номер записи: 48

02-01-2020 дата публикации

METHOD AND SYSTEM FOR SEQUENCING NUCLEIC ACIDS

Номер: US20200002762A1

Автор: BERNARD Mark A., DAMBACHER Corey M., ROKICKI Joseph, Tu Eugene, Vijayan Kandaswamy, WILSON Kerry

Принадлежит:

Provided are compositions, methods and systems for determining the sequence of a template nucleic acid using a polymerase-based, sequencing-by-binding procedure. An examination step involves monitoring the interaction between a polymerase and template nucleic acid in the presence of one or more nucleotides. Identity of the next correct nucleotide in the sequence is determined without incorporation of any nucleotide into the structure of the primer by formation of a phosphodiester bond. An optional incorporation step can be used after the examination step to extend the primer by one or more nucleotides, thereby incrementing the template nucleotides that can be examined in a subsequent examination step. The sequencing-by-binding procedure does not require the use of labeled nucleotides or polymerases, but optionally can employ these reagents. 1. A method of determining the identity of the next correct nucleotide for a primed template nucleic acid , the method comprising:(a) contacting the primed template nucleic acid with a plurality of nucleotide mixtures in serial fashion while precluding incorporation of any nucleotide into the primer, wherein each of the nucleotide mixtures comprises a combination of nucleotides that is complementary to two, three or four different types of bases in the template nucleic acid, thereby forming stabilized ternary complexes that comprise the primed template nucleic acid molecule, a polymerase and a next correct nucleotide from each of the nucleotide mixtures; and(b) detecting the stabilized ternary complexes while precluding incorporation of any nucleotide into the primer, thereby determining the identity of the next correct nucleotide for the primed template nucleic acid.2. The method of claim 1 , wherein the plurality of nucleotide mixtures comprises a plurality of different nucleotide mixtures.3. The method of claim 2 , wherein the plurality of different nucleotide mixtures comprises four different nucleotide mixtures.4. The method ...

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Номер записи: 49

02-01-2020 дата публикации

METHODS AND SYSTEMS FOR PROCESSING POLYNUCLEOTIDES

Номер: US20200002763A1

Автор: "OKeeffe Christopher Joachim", BELGRADER Phillip, Bent Zachary, Bharadwaj Rajiv, Boutet Stephane Claude, Gopalan Vijay Kumar Sreenivasa, Harada Josephine, Hindson Christopher, Lenji Mohammad Rahimi, Lucero Michael Ybarra, McDermott Geoffrey, Meer Elliott, Mikkelsen Tarjei Sigurd, Pfeiffer Katherine, Price Andrew D., Ryvkin Paul, Saxonov Serge, Srinivas Niranjan, Stuelpnagel John R., Taylor Sarah, Terry Jessica Michele, Wheeler Tobias Daniel, Wu Indira, Ziraldo Solongo Batjargal

Принадлежит:

The present disclosure provides compositions, methods, systems, and devices for polynucleotide processing and analyte characterization. Such polynucleotide processing may be useful for a variety of applications, including analyte characterization by polynucleotide sequencing. The compositions, methods, systems, and devices disclosed herein generally describe barcoded oligonucleotides, which can be bound to a bead, such as a gel bead, useful for characterizing one or more analytes including, for example, protein (e.g., cell surface or intracellular proteins), genomic DNA, and RNA (e.g., mRNA or CRISPR guide RNAs). Also described herein, are barcoded labelling agents and oligonucleotide molecules useful for “tagging” analytes for characterization. 130-. (canceled)31. A method for analyzing a tissue sample comprising:(a) delivering a plurality of spatial oligonucleotides to a location in a tissue sample comprising cells, wherein a spatial oligonucleotide of the plurality of spatial oligonucleotides comprises (i) a spatial barcode sequence and (ii) a cell labeling agent configured to deliver the spatial oligonucleotide to a cell at the location in the tissue sample;(b) dissociating the tissue sample into a plurality of labeled cells, wherein a labeled cell of the plurality of labeled cells comprises: (i) the spatial oligonucleotide and (ii) a plurality of analytes;(c) partitioning the labeled cell and a plurality of cell barcode nucleic acid molecules into a partition, wherein each cell barcode nucleic acid molecule of the plurality of cell barcode nucleic acid molecules (i) comprises a common cell barcode sequence and (ii) is configured to couple to the spatial oligonucleotide and analytes of the plurality of analytes;(d) in the partition, generating (i) a spatial barcoded nucleic acid molecule comprising the spatial barcode sequence or complement thereof, (ii) a first barcoded nucleic acid molecule corresponding to a first analyte of the plurality of analytes, and ( ...

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Номер записи: 50

02-01-2020 дата публикации

METHODS AND SYSTEMS FOR PROCESSING POLYNUCLEOTIDES

Номер: US20200002764A1

Автор: "OKeeffe Christopher Joachim", BELGRADER Phillip, Bent Zachary, Bharadwaj Rajiv, Boutet Stephane Claude, Gopalan Vijay Kumar Sreenivasa, Harada Josephine, Hindson Christopher, Lenji Mohammad Rahimi, Lucero Michael Ybarra, McDermott Geoffrey, Meer Elliott, Mikkelsen Tarjei Sigurd, Pfeiffer Katherine, Price Andrew D., Ryvkin Paul, Saxonov Serge, Srinivas Niranjan, Stuelpnagel John R., Taylor Sarah, Terry Jessica Michele, Wheeler Tobias Daniel, Wu Indira, Ziraldo Solongo Batjargal

Принадлежит:

The present disclosure provides compositions, methods, systems, and devices for polynucleotide processing and analyte characterization. Such polynucleotide processing may be useful for a variety of applications, including analyte characterization by polynucleotide sequencing. The compositions, methods, systems, and devices disclosed herein generally describe barcoded oligonucleotides, which can be bound to a bead, such as a gel bead, useful for characterizing one or more analytes including, for example, protein (e.g., cell surface or intracellular proteins), genomic DNA, and RNA (e.g., mRNA or CRISPR guide RNAs). Also described herein, are barcoded labelling agents and oligonucleotide molecules useful for “tagging” analytes for characterization. 130-. (canceled)31. A method for analyzing a tissue sample comprising:(a) contacting a plurality of cells with a plurality of labelling molecules to generate a plurality of labelled cells, wherein said plurality of labelling molecules comprise a plurality of cell barcode sequences, and wherein a labelled cell of said plurality of labeled cells comprises (i) a cell barcode sequence that is different from cell barcode sequences of other labelled cells of said plurality of labelled cells and (ii) a plurality of analytes;(b) generating a plurality of partitions comprising said plurality of labelled cells and a plurality of partition nucleic acid barcode molecules, wherein said plurality of partition nucleic acid barcode molecules comprise a plurality of partition barcode sequences, wherein a partition of said plurality of partitions comprises a partition barcode sequence that is different from partition barcode sequences of other partitions of said plurality of partitions, wherein a partition nucleic acid barcode molecule of said plurality of partition nucleic acid barcode molecules is configured to couple to an analyte from said plurality of analytes, wherein said analyte comprises messenger ribonucleic acid (mRNA), and wherein ...

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Номер записи: 51

05-01-2017 дата публикации

Compositions and methods for screening t cells with antigens for specific populations

Номер: US20170003288A1

Автор: James R. Heath, Songming PENG

Принадлежит: California Institute of Technology CalTech

Compositions and methods for isolating patient-derived antigen-specific T cells include an antigen complex having a polynucleotide barcoded nanoparticle sorting agent complexed with a peptide-loaded streptavidin major histocompatability complex (MHC) tetramer, the barcoding technology allowing for high fidelity screening of a library of the antigen complexes to readily isolate and identify antigen-specific T cells.

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Номер записи: 52

07-01-2021 дата публикации

MODIFIED AGPASE LARGE SUBUNIT SEQUENCES AND METHODS FOR DETECTION OF PRECISE GENOME EDITS

Номер: US20210002658A1

Автор: Begemann Matthew, January Emma

Принадлежит: BENSON HILL, INC.

Compositions and methods for improving plant growth are provided herein. Polynucleotides encoding modified AGPase large subunit proteins, polypeptides encompassing modified AGPase large subunit proteins, methods of producing modified polynucleotides encoding modified AGPase large subunit proteins, and expression constructs for expressing genes of interest whose expression may improve agronomic properties including but not limited to crop yield, biotic and abiotic stress tolerance, and early vigor, plants comprising the polynucleotides, polypeptides, and expression constructs, and methods of producing plants comprising the modified AGPase large subunit genes of the invention or encoding the AGPase large subunit proteins of the invention are also provided. Compositions and methods for the ready detection of precise base changes are also provided herein. Compositions include primer pads as part of a repair donor template, and methods for the detection of precise base changes include PCR detection using one or more primers designed to anneal with a primer pad sequence. 2. The modified AGPase large subunit protein of wherein said AGPase large subunit protein shares at least 80% identity with SEQ ID NO:6 claim 1 , 8 claim 1 , 10 claim 1 , 12 claim 1 , 14 claim 1 , or 16 claim 1 , or is encoded by a polynucleotide that shares at least 70% identity with a sequence selected from the group consisting of SEQ ID NOs:5 claim 1 , 7 claim 1 , 9 claim 1 , 11 claim 1 , 13 claim 1 , 15 claim 1 , and 43-48.4. The modified AGPase large subunit protein of wherein said AGPase large subunit protein shares at least 80% identity with a sequence selected from the group consisting of SEQ ID NOs:49-54 claim 3 , or is encoded by a polynucleotide that shares at least 70% identity with a sequence selected from the group consisting of SEQ ID NOs:55-66.5. The modified AGPase large subunit protein of wherein said modified AGPase large subunit has at least 80% sequence identity to a sequence selected ...

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Номер записи: 53

03-01-2019 дата публикации

NUCLEOTIDE-SPECIFIC RECOGNITION SEQUENCES FOR DESIGNER TAL EFFECTORS

Номер: US20190002824A1

Автор: Cong Le, Zhang Feng

Принадлежит:

The invention relates to methods of altering expression of a genomic locus of interest or specifically targeting a genomic locus of interest in an animal cell, which may involve contacting the genomic locus with a non-naturally occurring or engineered composition that includes a deoxyribonucleic acid (DNA) binding polypeptide having a N-terminal capping region, a DNA binding domain comprising at least five or more Transcription activator-like effector (TALE) monomers and at least one or more half-monomers specifically ordered to target the genomic locus of interest, and a C-terminal capping region, wherein the polypeptide includes at least one or more effector domains, and wherein the polypeptide is encoded by and translated from a codon optimized nucleic acid molecule so that the polypeptide preferentially binds to the DNA of the genomic locus. 110-. (canceled)11. A non-naturally occurring or engineered composition comprising a deoxyribonucleic acid (DNA) binding polypeptide comprising:(a) a N-terminal capping region,(b) a DNA binding domain comprising at least five or more Transcription activator-like effector (TALE) monomers and at least one or more half-monomers specifically ordered to target a genomic locus of interest, and(c) a C-terminal capping region,wherein (a), (b) and (c) are arranged in a predetermined N-terminus to C-terminus orientation,wherein the polypeptide includes at least one or more mSin interaction domain (SID) repressor domain having the sequence MNIQMLLEAADYLERREREAEHGYASMLP (SEQ ID NO: 49),{'sub': 1-11', '12', '13', '14-33 or 34 or 35', 'z, 'wherein the DNA binding domain comprises (X-XX-X),'}{'sub': '1-11', 'wherein Xis a chain of 11 contiguous amino acids,'}{'sub': 12', '13, 'wherein XXis a repeat variable diresidue (RVD),'}{'sub': '14-33 or 34 or 35', 'wherein Xis a chain of 21, 22 or 23 contiguous amino acids,'}wherein z is at least 5 to 40, and{'sub': '13', 'wherein at least one RVD is selected from the group consisting of HH, KH, NH, ...

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Номер записи: 54

03-01-2019 дата публикации

RNase H-Based Assays Utilizing Modified RNA Monomers

Номер: US20190002865A1

Автор: BEHLKE Mark Aaron, DOBOSY Joseph, Rose Scott, WALDER Joseph Alan

Принадлежит:

The present invention pertains to novel oligonucleotide compounds for use in various biological assays, such as nucleic acid amplification, ligation and sequencing reactions. The novel oligonucleotides comprise a ribonucleic acid domain and a blocking group at or near the 3′ end of the oligonucleotide. These compounds offer an added level of specificity previously unseen. Methods for performing nucleic acid amplification, ligation and sequencing are also provided. Additionally, kits containing the oligonucleotides are also disclosed herein. 1. A kit for the ligation of the donor and accepter ends of a single oligonucleotide in the presence of a target nucleic acid sequence comprising:(a) an oligonucleotide in which either its donor end, its accepter end, or both its donor and accepter ends comprise a cleavage domain and a blocking group preventing ligation of its 5′ phosphate group to its 3′ OH group;(b) a cleaving agent;(c) a ligation agent; and(d) optionally, an instruction manual for ligating the accepter and donor ends of the oligonucleotide in the presence of a target nucleic acid sequence.2. The kit of claim 1 , wherein the cleavage domain is an RNase H cleavable domain.3. The kit of claim 2 , wherein the cleavage domain comprises a single RNA residue.4. The kit of claim 2 , wherein the cleavage domain lacks an RNA residue.5. The kit of claim 4 , wherein the cleavage domain comprises one or more 2′-modified nucleosides.6. The kit of claim 1 , wherein the cleaving agent is an RNase H enzyme.7. The kit of claim 6 , wherein the RNase H enzyme is an RNase H2 enzyme.8. The kit of claim 1 , wherein the ligation agent is a DNA ligase enzyme.9. The kit of claim 8 , wherein the DNA ligase enzyme is a thermostable DNA ligase.10. The kit of claim 9 , wherein the thermostable DNA ligase is a hot-start DNA ligase having reduced activity at lower temperatures.11. The kit of claim 10 , wherein the DNA ligase inherently has lower activity at reduced temperatures or is ...

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Номер записи: 55

03-01-2019 дата публикации

METHOD AND COMPOSITION

Номер: US20190002870A1

Автор: Rogers Jan, TORO RUEDA Carlos

Принадлежит:

A method of extracting DNA and/or RNA from a cell or capsid, the method comprising steps of: (a) contacting a composition comprising the cell or capsid with a composition comprising (ii) a quaternary ammonium compound or a precursor thereof; and (b) contacting the composition obtained in step (a) with a composition comprising a proteinaceous washing agent. 1. A method of extracting DNA and/or RNA from a cell or capsid , the method comprising steps of: '(i) a quaternary ammonium compound or a precursor thereof; and', '(a) contacting a composition comprising the cell or capsid with a composition comprising'}(b) contacting the composition obtained in step (a) with a composition comprising a proteinaceous washing agent.2. A method according to wherein the composition comprising the cell or capsid is a blood sample.3. A method according to wherein the composition comprising the cell or capsid is a whole blood sample.6. A method according to wherein the composition contacted with the cell or capsid in step (a) further comprises (ii) a non-ionic surfactant.7. A method according to wherein the non-ionic surfactant is an alkyl polyglucoside (APG).9. A method according to wherein the composition contacted with the composition comprising the cell or capsid in step (a) has a pH of from 6 to 8.10. A method according to wherein there are no purification steps between step (a) and step (b).11. A method according to wherein the proteinaceous washing agent is selected from bovine serum albumin and casein.12. A kit comprising: (i) a quaternary ammonium; and optionally', '(ii) a non-ionic surfactant; and, '(a) a first composition comprising'}(b) a second composition comprising an proteinaceous washing agent.13. A method of identifying a component of genetic material claim 1 , the method comprising the steps of: (i) a quaternary ammonium compound; and optionally', '(ii) a non-ionic surfactant;, '(a) contacting a composition comprising a cell or capsid with a composition comprising(b) ...

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Номер записи: 56

07-01-2021 дата публикации

BACTERIAL IDENTIFICATION METHOD USING RNA OF SAMPLE BACTERIA, AND KIT THEREFOR

Номер: US20210002709A1

Автор: AMANO Koh, ENDO Ayako, Morishige Takashi, TSUJI Kentaro, YANAI Hisaaki

Принадлежит: Mitsui Chemicals, Inc.

A method for identifying a bacterium includes: (1) A step of performing a reverse transcription reaction using a first reaction system consisting of a primer for reverse transcription to prepare a cDNA containing a base sequence for identification of a bacterium of interest, an RNA extracted from the bacterium in a sample, and an enzyme with RNA-dependent DNA polymerase activity to obtain a reaction mixture containing the synthesized cDNA and the primers for the reverse transcription, (2) A step of performing PCR using a second reaction system consisting of the reaction mixture, a primer pair for synthesis of a double-stranded DNA containing the base sequence for identification of the bacterium of interest, and an enzyme with DNA-dependent DNA polymerase activity, and (3) A step of detecting the generation of the double-stranded DNA containing the base sequence for identification of the bacterium of interest from the second reaction system after PCR. 1. A bacterial identification method having the following steps (1) to (3):(1) A step of performing a reverse transcription reaction using a first reaction system comprising a primer for reverse transcription to prepare a cDNA containing a base sequence for identification of a bacterium of interest, an RNA extracted from the bacterium in a sample, and an enzyme with RNA-dependent DNA polymerase activity to obtain a reaction mixture containing the synthesized cDNA and the primers for the reverse transcription,(2) A step of performing PCR using a second reaction system comprising the reaction mixture, a primer pair for synthesis of a double-stranded DNA containing the base sequence for identification of the bacterium of interest, and an enzyme with DNA-dependent DNA polymerase activity, provided that in the second reaction system, the concentration of the primer for reverse transcription supplied from the reaction mixture is not less than 0.08 nM and not more than 20 nM, and(3) A step of detecting the generation of the ...

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Номер записи: 57

07-01-2021 дата публикации

QUANTITATIVE MICROBIAL COMMUNITY PROFILING USING MOLECULAR INVERSION PROBES WITH UNIQUE MOLECULAR IDENTIFIERS

Номер: US20210002718A1

Автор: HARTMAN Wyatt, Zwirko Zachary

Принадлежит: Trace Genomics, Inc.

The present disclosure relates to the profiling of microorganisms in an environmental sample. Specifically, the present disclosure relates to methods of using molecular inversion probes comprising unique molecular identifiers to profile microorganisms in an environmental sample. In addition, the present disclosure relates to compositions of molecular inversion probes comprising unique molecular identifiers to profile microorganisms in an environmental sample. 1. A method for profiling of microorganisms in an environmental sample , wherein the method comprisesa) extracting DNA from the environmental sample;b) denaturing the extracted DNA; wherein the MIP comprises', '(i) in the 3′ to 5′ direction,', 'a first target locus primer, wherein the first primer comprises a nucleotide sequence complementary to a first sequence in a target locus,', 'a universal backbone sequence comprising a first sequencing primer binding site and a second sequencing primer binding site, and', 'a second target locus primer, wherein the second primer comprises a nucleotide sequence complementary to a second, non-overlapping sequence in the target locus, and', '(ii) a first unique molecular identifier (UMI);', 'wherein the backbone sequence has low sequence homology to DNA in the environmental sample and has minimal ability to form secondary structures,', 'thereby generating a sample comprising denatured DNA-MIP complexes;, 'c) incubating the denatured DNA with a molecular inversion probe (MIP) under conditions that allow hybridization,'}d) after hybridization, performing an extension and ligation reaction comprising incubating the sample comprising denatured DNA-MIP complexes with nucleotides, 5′ exo-polymerase lacking strand displacement activity, and a thermostable ligase capable of ligating splinted substrates under conditions that allow extension of the 3′ end of the MIP and ligation to the 5′ end of the MIP;e) after extension and ligation, incubating the sample comprising denatured DNA- ...

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Номер записи: 58

03-01-2019 дата публикации

A novel method for the preparation of bar-coded primer sets

Номер: US20190002953A1

Автор: Can SÖNMEZER, Christian KRENDL, Florian OPPERER, Micha Drukker

Принадлежит: Helmholtz Zentrum Muenchen Deutsches Forschungszentrum fuer Gesundheit und Umwelt GmbH, Helmholtz Zentrum Muenchendeutsches Forschungszentrum fur Gesundheit und Umwelt

The present invention relates to a method of producing a set of primers suitable for the reverse transcription and/or amplification of a plurality (N) of nucleic acid molecules of interest, wherein for each nucleic acid molecule of interest at least one primer is produced and wherein the primers carry a bar-code, the method comprising the steps of: (a)(i) combining (1) a first oligonucleotide, wherein said first oligonucleotide comprises a first bar-code nucleic acid sequence linked at its 3′ end to a first adapter nucleic acid sequence with (2) a plurality (N) of second oligonucleotides, wherein each second oligonucleotide comprises the reverse complementary sequence of a forward primer specific for a nucleic acid molecule of interest, wherein said reverse complementary sequence of the forward primer is linked at its 3′ end to the reverse complementary sequence of the first adapter nucleic acid sequence; and/or (a)(ii) combining (1) a third oligonucleotide, wherein said third oligonucleotide comprises a second bar-code nucleic acid sequence linked at its 3′ end to a second adapter nucleic acid sequence with (2) a plurality (N) of fourth oligonucleotides, wherein each fourth oligonucleotide comprises the reverse complementary sequence of a reverse primer specific for said nucleic acid molecule of interest, wherein said reverse complementary sequence of the reverse primer is linked at its 3′ end to the reverse complementary sequence of the second adapter nucleic acid sequence; wherein steps (a)(i) and (a)(ii) are carried out under conditions that enable the annealing of the first and second adapter nucleic acid sequences to the respective reverse complementary sequences thereof; (b) extending the oligonucleotides of (a)(i) and (a)(ii) by polymerase-mediated oligonucleotide synthesis; and (c) optionally, removing the second and fourth oligonucleotides. The present invention further relates to methods of producing a plurality (M) of nucleic acid amplification products ...

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Номер записи: 59

03-01-2019 дата публикации

NUCLEIC ACID AMPLIFICATION

Номер: US20190002957A1

Автор: Erlander Mark G., SALUNGA Ranelle C.

Принадлежит:

The present invention provides methods for the amplification of nucleic acid molecules. Methods for amplifying target polynucleotides, including mRNA, using oligonucleotides, DNA and RNA polymerases are provided. The invention further provides compositions and kits for practicing the methods, as well as methods which use the amplification products. 120.-. (canceled)21. A method of amplifying RNA sequences , comprising:annealing a single-stranded target polynucleotide with a first oligonucleotide that includes a primer region comprising an oligo dT sequence of about 18-21 T residues and a promoter region comprising a T7 promoter sequence to form a first complex;synthesizing a first strand cDNA by reverse transcription of the first complex to form an RNA/cDNA heteroduplex;separating the first strand cDNA from the RNA/cDNA heteroduplex;annealing the first strand cDNA with a plurality of random primers that hybridize at a plurality of positions on the first strand cDNA to form a second complex;forming a double-stranded cDNA template from the second complex using a combination of DNA dependent polymerase enzymes including exonuclease deficient Klenow and Taq polymerase; andtranscribing the double-stranded cDNA template with an RNA polymerase capable of initiating transcription via said promoter region to produce amplified RNA (aRNA) containing a sequence complementary to the single-stranded target polynucleotide.22. The method of claim 21 , wherein the first oligonucleotide further comprises an anchor sequence between the primer region and the promoter region.23. The method of claim 21 , further comprising degrading the first oligonucleotide remaining after synthesizing a first strand cDNA with an exonuclease.24. The method of claim 23 , wherein the exonuclease is exonuclease I.25. The method of claim 21 , wherein separating the first strand cDNA from the RNA/cDNA heteroduplex comprises denaturing the RNA/cDNA heteroduplex.26. The method of claim 25 , wherein denaturing ...

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Номер записи: 60

03-01-2019 дата публикации

GENETIC TESTING FOR ALIGNMENT-FREE PREDICTING RESISTANCE OF MICROORGANISMS AGAINST ANTIMICROBIAL AGENTS

Номер: US20190002960A1

Автор: Backes Christina, GALATA VALENTINA, Keller Andreas, Schmolke Susanne, Stähler Cord Friedrich

Принадлежит:

The present invention relates to a method of determining an infection of a patient with at least one microorganism, particularly a bacterial microorganism, potentially resistant to antimicrobial drug treatment, a method of selecting a treatment of a patient suffering from an infection with at least one microorganism, particularly bacterial microorganism, and a method of determining an antimicrobial drug, e.g. antibiotic, resistance profile for at least one microorganism, particularly bacterial microorganism, as well as computer program products used in these methods. 1. A method of determining an antimicrobial drug , e.g. antibiotic , resistance profile for a microorganism , particularly a bacterial microorganism , comprising:obtaining or providing a first data set of gene sequences of a plurality of clinical isolates of the microorganism;wherein at least a part of the gene sequences of the first data set are assembled;analyzing the gene sequences of the first data set for genetic variants to obtain a third data set of genetic variants;providing a second data set of antimicrobial drug, e.g. antibiotic, resistance and/or susceptibility of the plurality of clinical isolates of the microorganism;correlating the third data set with the second data set and statistically analyzing the correlation; anddetermining the genetic sites in the genome of the microorganism with antimicrobial drug, e.g. antibiotic, resistance.2. The method of claim 1 , wherein the genetic variants in the gene sequences of the first data set are single nucleotide polymorphisms (SNPs).3. The method of claim 2 , wherein the SNPs are detected alignment-free.4. The method of or claim 2 , wherein the SNPs are annotated to a pan-genome of the microorganism and/or annotated to one or more reference genomes.5StaphylococcusStaphylococcus aureus,. The method of any one of the preceding claims claim 2 , wherein the microorganism is a species claim 2 , particularly optionally wherein the antimicrobial drug is ...

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Номер записи: 61

03-01-2019 дата публикации

BODILY FLUID TARGET ENRICHMENT

Номер: US20190002964A1

Автор: Shuber Anthony P.

Принадлежит:

The invention provides methods for capturing target nucleic acid directly from bodily fluid samples, without the need for certain complex sample preparation steps, using Cas endonuclease to bind to the target nucleic acid sequences. The Cas proteins, along with their sequence-specific guide RNAs, may be introduced directly into the sample, where the Cas proteins bind to ends of a target nucleic acid. The target nucleic acid is thus isolated or enriched in a sequence-specific manner. The target nucleic acid may then be subject to any suitable detection or analysis assay, such as amplification or sequencing. The target nucleic acid may be enriched by digesting other, unbound nucleic acids present in the sample with exonuclease. The bound Cas proteins prevent exonuclease from digesting the target nucleic acid, thereby leaving the only the target nucleic acid substantially present in the sample. 1. A method for enriching a sample , the method comprising:obtaining a bodily fluid sample comprising a target nucleic acid; andintroducing Cas endonuclease to the bodily fluid sample to bind to the target nucleic acid.2. The method of claim 1 , wherein the Cas endonuclease is a catalytically inactive homolog thereof.3. The method of claim 2 , wherein the introduction step comprises introducing the Case endonuclease and guide RNA into the bodily fluid sample and binding the Cas endonuclease to ends of the target nucleic acid.4. The method of claim 1 , further comprising:introducing an exonuclease to the bodily fluid sample to digest unbound nucleic acid.5. The method of claim 1 , wherein the bodily fluid sample comprises bile claim 1 , blood claim 1 , plasma claim 1 , serum claim 1 , sweat claim 1 , saliva claim 1 , urine claim 1 , feces claim 1 , phlegm claim 1 , mucus claim 1 , sputum claim 1 , tears claim 1 , cerebrospinal fluid claim 1 , synovial fluid claim 1 , pericardial fluid claim 1 , lymphatic fluid claim 1 , semen claim 1 , vaginal secretion claim 1 , products of ...

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Номер записи: 62

03-01-2019 дата публикации

POLYPEPTIDE TAGGED NUCLEOTIDES AND USE THEREOF IN NUCLEIC ACID SEQUENCING BY NANOPORE DETECTION

Номер: US20190002968A1

Автор: Bergmann Frank, Gremyachinskiy Dmitriy, HILLRINGHAUS Lars, KUCHELMEISTER Hannes, SEIDEL Christoph, TRANS Andrew

Принадлежит:

The present disclosure relates to compositions and methods based on polypeptide-tagged nucleotide, and the use of such polypeptide-tagged nucleotides in nanopore devices and methods. 1. A compound of structural formula (I){'br': None, 'N-P-L-T\u2003\u2003 (I)'} N is a nucleoside;', 'P is an oligophosphate covalently attached to a 5′-O group of the nucleoside,', 'wherein the oligophosphate consists of 3 to 12 phosphate groups;', 'L is a linker covalently attached to a terminal phosphate group of the oligophosphate; and', 'T is a polypeptide tag covalently attached to the linker, wherein the polypeptide has an overall charge and comprises at least one helical structure., 'wherein,'}3. The compound of claim 1 , wherein the length of the polypeptide tag is at least 16 amino acid residues claim 1 , optionally wherein the length of the polypeptide tag is at least 20 amino acid residues claim 1 , at least 25 amino acid residues claim 1 , at least 30 amino acid residues claim 1 , at least 40 amino acid residues claim 1 , at least 50 amino acid residues claim 1 , at least 60 amino acid residues claim 1 , at least 70 amino acid residues claim 1 , at least 80 amino acid residues claim 1 , or at least 90 amino acid residues.4. The compound of claim 1 , wherein the helical structure comprises is at least 8 amino acid residues claim 1 , optionally wherein the polypeptide helical structure comprises at least 16 amino acid residues claim 1 , at least 20 amino acid residues claim 1 , at least 25 amino acid residues claim 1 , at least 30 amino acid residues claim 1 , at least 40 amino acid residues claim 1 , at least 50 amino acid residues claim 1 , or at least 60 amino acid residues.5. The compound of claim 1 , wherein the helical structure is an α-helix claim 1 , optionally wherein the length of the α-helix is at least 10 amino acid residues claim 1 , at least 16 amino acid residues claim 1 , at least 20 amino acid residues claim 1 , at least 25 amino acid residues claim 1 , at ...

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Номер записи: 63

03-01-2019 дата публикации

ENZYME-PORE CONSTRUCTS

Номер: US20190002972A1

Автор: Bayley John Hagan Pryce, CHELEY Stephen, Clarke James Anthony, Jayasinghe Lakmal, MCKEOWN Brian, White James

Принадлежит: Oxford Nanopore Technologies Ltd.

The invention relates to constructs comprising a transmembrane protein pore subunit and a nucleic acid handling enzyme. The pore subunit is covalently attached to the enzyme such that both the subunit and enzyme retain their activity. The constructs can be used to generate transmembrane protein pores having a nucleic acid handling enzyme attached thereto. Such pores are particularly useful for sequencing nucleic acids. The enzyme handles the nucleic acid in such a way that the pore can detect its component nucleotides by stochastic sensing. 153-. (canceled)54. A method of attaching an enzyme to a transmembrane protein pore subunit comprising:(a) providing an enzyme with a first linker wherein one end of the first linker is attached to the carboxyl or amino terminus of the enzyme and the other end of the first linker is provided with a reactive group;(b) providing a transmembrane protein pore subunit with a second linker wherein one end of the second linker is attached to the carboxyl or amino terminus of the protein pore subunit and the other end of the second linker is provided with a reactive group that is specific for the reactive group in (a);(c) contacting the enzyme and protein pore subunit of (a) and (b) above under suitable conditions such that the reactive group of the first linker attaches to the reactive group of the second linker, thereby attaching the enzyme and transmembrane protein pore subunit.55. A method according to wherein the first linker is attached to the carboxyl terminus of the enzyme.56. A method according to wherein the second linker is attached to the carboxyl terminus of the protein pore subunit.57. A method according to wherein the first linker is attached to the carboxyl terminus of the enzyme and the second linker is attached to the carboxyl terminus of the protein pore subunit.58. A method according to wherein the reactive group of the first linker forms a covalent bond with the reactive group of the second linker claim 54 , such ...

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Номер записи: 64

03-01-2019 дата публикации

APPARATUS AND METHODS FOR KINETIC ANALYSIS AND DETERMINATION OF NUCLEIC ACID SEQUENCES

Номер: US20190002973A1

Автор: Chen Cheng-Yao, He Molly, Pantoja Rigo, Previte Michael, Zhou Chunhong

Принадлежит: Illumina, Inc.

A method of distinguishing nucleotide sequences for different nucleic acid molecules including the steps of (a) mixing a plurality of different nucleic acid molecules with polymerase molecules and nucleotide molecules, wherein the different nucleic acid molecules are attached to a surface in the form of an array of nucleic acid features; (b) determining a transient state of the polymerase molecules at the nucleic acid features; and (c) identifying a subset of nucleic acid features that correctly incorporate the nucleotide molecules based on the transient state of the polymerase molecules at the nucleic acid features, thereby distinguishing the nucleotide sequences for the different nucleic acid molecules. 1. A method of distinguishing nucleotide sequences for different nucleic acid molecules , comprising(a) mixing a plurality of different nucleic acid molecules with polymerase molecules and nucleotide molecules,wherein the different nucleic acid molecules are attached to a surface in the form of an array of nucleic acid features;(b) monitoring binding of the polymerase molecules to the nucleic acid features at several points during a pre-equilibrium time period, thereby determining a transient state of the polymerase molecules at the nucleic acid features; and(c) identifying nucleic acid features of the array that correctly incorporate the nucleotide molecules based on the transient state of the polymerase molecules at the nucleic acid features, thereby distinguishing the nucleotide sequences for the different nucleic acid molecules.2. The method of claim 1 , wherein (b) comprises monitoring pre-steady state kinetic rate profiles of the binding of the polymerase molecules to the nucleic acid features.3. The method of claim 2 , wherein the monitoring of the pre-steady state kinetic rate profiles occurs before claim 2 , during claim 2 , and after correct incorporation of the nucleotide molecules into the nucleic acid features.4. The method of claim 2 , wherein (c) ...

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Номер записи: 65

01-01-2015 дата публикации

METHOD FOR SUPPRESSING THE EFFECTS OF ASCORBIC ACID

Номер: US20150004635A1

Автор: Soya Haruyo

Принадлежит:

Provided is a method for suppressing an effect of ascorbic acid on a measurement of glycated hemoglobin in a glycated hemoglobin-containing sample. The method of the present invention is a method for suppressing an effect of ascorbic acid on the measurement of glycated hemoglobin in a glycated hemoglobin-containing sample, comprising reacting the glycated hemoglobin-containing sample with a proteolytic enzyme, reacting a generated glycated peptide with a glycated peptide oxidase, and then measuring a generated hydrogen peroxide, wherein the reaction of the glycated peptide with the glycated peptide oxidase is performed in the presence of a halogen oxide and a substance selected from the group consisting of a substance represented by formula (I), wherein Rrepresents substituted or unsubstituted alkyl and a substance represented by the general formula (II), wherein Rrepresents substituted or unsubstituted alkyl; and represents a single bond or a double bond. 2. The method according to claim 1 , wherein the halogen oxide is selected from the group consisting of iodic acid or a salt thereof and bromic acid or a salt thereof.3. The method according to claim 1 , wherein the glycated hemoglobin is hemoglobin A1c.5. The reagent according to claim 4 , wherein the halogen oxide is selected from the group consisting of iodic acid or a salt thereof and bromic acid or a salt thereof.6. The reagent according to claim 4 , wherein the glycated hemoglobin is hemoglobin A1c.7. The reagent according to claim 4 , wherein the reagent suppresses an effect of ascorbic acid on the measurement of glycated hemoglobin in the sample.8. The method according to claim 2 , wherein the glycated hemoglobin is hemoglobin A1c.9. The reagent according to claim 5 , wherein the glycated hemoglobin is hemoglobin A1c.10. The reagent according to claim 5 , wherein the reagent suppresses an effect of ascorbic acid on the measurement of glycated hemoglobin in the sample.11. The reagent according to claim 6 , ...

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Номер записи: 66

04-01-2018 дата публикации

Methods and kits for measuring and quantifying dna double-stranded breaks using gamma-h2ax and h2ax

Номер: US20180003720A1

Автор: Christophe E. Redon, Jiuping Ji, William M. Bonner, Yiping Zhang

Принадлежит: US Department of Health and Human Services

Disclosed are methods of measuring the amount of exposure of a host to a DNA double-stranded break (DSB)-causing agent by determining the ratio of the quantity of γ-H2AX to the quantity of total H2AX in a biological sample from the host as compared to the ratio of the quantity of γ-H2AX to the quantity of total H2AX in a positive control biological sample that has been exposed to a known amount of a DSB-causing agent. Related kits and methods of quantifying DSBs in a test biological sample are also disclosed.

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Номер записи: 67

02-01-2020 дата публикации

Method and system for substance detection with a magnetic sensor

Номер: US20200003767A1

Автор: Jiaoming Qiu

Принадлежит: Zepto Life Technology Inc

Methods, systems and programing for substance detection with a magnetic sensor are presented. In one example, a magnetic sensor having one or more layers is formed on a base for sensing a magnetic field created by magnetic particles present in proximity to the magnetic sensor. A first end of each of a first set of strands is immobilized with respect to the magnetic sensor. A magnetic particle is attached to a second end of each of the first set of strands so that when a material containing a substance is in contact with the base, the substance causes at least some of the first set of strands to break resulting in that the magnetic particle attached to the second end of each of the at least some of the first set of strands is no longer in proximity to the magnetic sensor.

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Номер записи: 68

03-01-2019 дата публикации

Optical analyte detection systems and methods of use

Номер: US20190003975A1

Автор: Abraham J. Qavi, Adam L. Washburn, Jared T. Kindt, Ji-Yeon Byeon, Lawrence C. Gunn, III, Martin Anthony Gleeson, Matthew S. Luchansky, Melinda S. Mcclellan, Ryan C. Bailey, Tate Owen

Принадлежит: GENALYTE Inc, University of Illinois

Various embodiments are drawn to systems and methods for detecting an analyte of interest in a sample including an optical sensor, a capture probe attached to a surface of the optical sensor wherein the capture probe is capable of binding to the analyte to form a duplex or complex, and an antibody capable of binding to the analyte, duplex, or complex. In several embodiments, systems and methods further include a particle attached to the antibody or capable of binding to the antibody. In several embodiments, systems and methods for analyte detection feature one or more of the following: high detection sensitivity and specificity, scalability and multiplex capacity, ability to analyze large analytes, and ability to detect or measure multiple individual binding events in real-time.

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Номер записи: 69

03-01-2019 дата публикации

METHOD AND APPARATUS FOR ANALYTE DETECTION USING AN ELECTROCHEMICAL BIOSENSOR

Номер: US20190004005A1

Автор: Feldman Benjamin, Oja Stephen M.

Принадлежит:

A method for sensing an analyte utilizing a sensor having a working electrode, the method includes providing the working electrode with an analyte-specific enzyme and a redox mediator, providing the working electrode to the analyte, accumulating charge derived from the analyte reacting with the analyte-specific enzyme and the redox mediator for a set period of time, connecting the working electrode to circuit after the set period of time, and measuring the signal from the accumulated charge. 1. A method for sensing an analyte utilizing a sensor , the sensor including a working electrode , the method comprising:providing the working electrode with an analyte-specific enzyme and a redox mediator;providing the working electrode to the analyte;accumulating charge derived from the analyte reacting with the analyte-specific enzyme and the redox mediator for a set period of time;connecting the working electrode to a circuit after the set period of time; andmeasuring a signal from the accumulated charge.2. The method of claim 1 , wherein prior to providing the working electrode to an analyte claim 1 , the method further comprises connecting the working electrode to the circuit claim 1 , and prior to providing the working electrode to the analyte claim 1 , the method further comprises disconnecting the working electrode from the circuit.3. The method of claim 1 , wherein the working electrode is connected to the circuit prior to providing the working electrode to the analyte claim 1 , the method further comprises disconnecting the working electrode from the circuit prior to providing the working electrode to the analyte.4. The method of claim 1 , wherein the sensor is an enzymatic electrochemical biosensor.5. The method of claim 1 , wherein the redox mediator is an immobilized redox polymer.6. The method of claim 1 , wherein the analyte is selected from the group consisting of cortisol claim 1 , glucose claim 1 , lactate claim 1 , 3-hydroxy butyrate claim 1 , alcohol claim 1 ...

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Номер записи: 70

03-01-2019 дата публикации

Design, synthesis and use of synthetic nucleotides comprising charge mass tags

Номер: US20190004055A1

Автор: Jonathan J. O'Halloran, Joseph H. HEDLEY

Принадлежит: QUANTUMDX GROUP Ltd

Embodiments of the present disclosure relate generally to reporter compositions which are synthetic nucleotides that comprise nucleotides with a high charge mass moiety attached thereto via a linker molecule. The linker molecules can vary in length in part to enable the high charge mass moiety to extend out from a DNA polymerase complex so that polymerization may not be influenced.

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Номер записи: 71

03-01-2019 дата публикации

CONTROL OF pH IN AQUEOUS UREA-CONTAINING SOLUTIONS UTILIZING AMINO ACID-CONTAINING COMPOSITIONS

Номер: US20190004072A1

Автор: Horan Kevin, Uretsky Laura

Принадлежит: SIEMENS HEALTHCARE DIAGNOSTICS INC.

Aqueous calibration or quality control reagents that include urea are disclosed; the reagents may further include at least one amino acid-containing composition to provide pH stability thereto. Methods of production and use thereof are also disclosed. 1. A composition comprising an aqueous diagnostic quality control or calibration reagent disposed in a closed system , the composition comprising:urea present in a concentration in a range of from about 0.1 mM to about 150 mM;at least one buffer; andat least one amino acid-containing composition having at least one available primary or secondary amine, the at least one amino acid-containing composition consisting of ornithine, wherein a ratio of urea to the at least one amino acid-containing composition is in a range of from about 0.1:1 to about 100:1, and wherein the pH of the aqueous quality control or calibration reagent varies by +/−1.0 or less of the original pH following storage of the composition, and further wherein formation of ammonia in the aqueous diagnostic quality control or calibration reagent is substantially reduced by at least 5% following storage of the composition when compared to a composition prepared in the absence of the at least one amino acid-containing composition and following storage thereof; andwherein the aqueous diagnostic quality control or calibration reagent is for utilization with a diagnostic sensor.2. The composition of claim 1 , wherein the formation of ammonia in the aqueous diagnostic quality control or calibration reagent is substantially reduced by at least 10% when compared to a composition prepared in the absence of the at least one amino acid-containing composition following storage thereof.3. The composition of claim 1 , wherein the formation of ammonia in the aqueous diagnostic quality control or calibration reagent is substantially reduced by at least 15% when compared to a composition prepared in the absence of the at least one amino acid-containing composition ...

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Номер записи: 72

03-01-2019 дата публикации

DIGITAL MEASUREMENTS FROM TARGETED SEQUENCING

Номер: US20190005193A1

Автор: Amorese Douglas A., HUELGA Stephanie C., SCHROEDER Benjamin G., Scolnick Jonathan

Принадлежит:

Disclosed herein are methods, compositions and kits for quantitating one or more specific nucleic acids within a plurality of nucleic acids. In some embodiments, a sequencing library is constructed from enriched probe extension products specific for the specific nucleic acids and sequenced. In some embodiments, the resulting reads are used for removing duplicate reads. In some embodiments, counting of verified probes is used to quantitate or determine the number of specific nucleic acid molecules in the starting nucleic acid sample. 1. A method for quantitating a plurality of specific nucleic acid molecules in a composition comprising: a. generating a plurality of probe extension products , wherein each probe extension product comprises a probe sequence that is complementary to a probe target region within a specific nucleic acid molecule; b. sequencing the plurality of probe extension products to generate a sequence for each of the plurality of probe extension products; c. aligning the sequence of each of the plurality of probe extension products to a reference sequence database , wherein the reference sequence database comprises probe sequences; and d. determining the number of alignments for the sequence of each probe extension product with a sequence in the reference sequence database , wherein the number of alignments indicates the quantity of each of the specific nucleic acid molecule that the probe of the probe extension product is complementary to.2. A method for quantitating a plurality of specific nucleic acid molecules comprising: a. generating a plurality of probe extension products , wherein each probe extension product comprises (i) a first adapter , and (ii) a probe sequence complementary to a probe target region within a specific nucleic acid molecule; b. sequencing the plurality of probe extension products to generate sequence data comprising a sequence for each of the plurality of probe extension products; c. identifying the presence of the probe ...

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Номер записи: 73

03-01-2019 дата публикации

Cancer evolution detection and diagnostic

Номер: US20190005194A1

Автор: AmirAli TALASAZ, Helmy Eltoukhy

Принадлежит: Guardant Health Inc

The present disclosure provides methods for determining a probability that after any of a number of therapeutic interventions, an initial state of a subject, such as somatic cell mutational status of a subject with cancer, will develop a subsequent state. Such probabilities can be used to inform a health care provider as to particular courses of treatment to maximize probability of a desired outcome for the subject.

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Номер записи: 74

14-01-2021 дата публикации

IDENTIFICATION OF WHITE LEGHORNS RED PLUMAGE MUTAGENIC MUTANT GENOTYPE AND CULTIVATION METHOD FOR SUPPORTING SYSTEM OF RED PLUMAGE PINK SHELL LAYER CHICKENS

Номер: US20210007334A1

Автор: Li Guangqi, Li Huani, Liu Aiqiao, Sun Congjiao, SUN Hao, Wu Guiqin, Yang Ning

Принадлежит:

The present invention discloses a method for breeding the commercial strains of red feather pink-shell laying hens. It provides a primer pair for identifying the red feather causative mutation homozygous genotype of white leghorn chickens, which is composed of the single-stranded DNA molecule shown in Sequence 2 of the Sequence List and the single-stranded DNA molecule shown in Sequence 3 of the list. After the primer was designed according to the upstream and downstream nucleotide sequences of the 18,288,303deoxynucleotide in the positive-sense strand of the 11chromosome as shown in the sequence information of the chicken reference genome Gallus_gallus-4.0 version published in NCBI, the genotype (SNP) at this site is tested through the restriction fragment length polymorphism, the genotype of the site (SNP) was tested through the restriction fragment length polymorphism; the offspring hens obtained by cross breeding the homogenous female parent (the homogenous female parent was obtained through expanded propagation of the white leghorn chickens with the red feather causative mutation homozygous genotype) and the Rhode Island Red rooster as a male parent are all of red feather phenotype, meeting the market demands and enjoying a broad prospect for promotion. 1. A primer pair for identifying the red feature causative mutation genotypes of white leghorn chickens , which is composed of the single-stranded DNA molecule shown in Sequence 2 of the sequence list and the single-stranded DNA molecule shown in Sequence 3 of the list.2. A kit for identifying the red feature causative mutation genotypes of white leghorn chickens claim 1 , consisting of the primer pair and the restriction endonuclease according to ;and the said restriction endonuclease is NruI.3. The application of the primer pair according to in identifying the genotypes of the red feather causative mutation of the white leghorn chickens to be tested.4. The application of the primer pair according to in the ...

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Номер записи: 75

11-01-2018 дата публикации

IMPROVED DROPLET SEQUENCING APPARATUS AND METHOD

Номер: US20180008985A1

Автор: BALMFORTH Barnaby, Frayling Cameron Alexander, ISAAC Thomas Henry

Принадлежит: BASE4 INNOVATION LTD

An apparatus for sequencing a polynucleotide analyte is provided and comprises; •a first zone in which a stream of single nucleotides is generated by progressive digestion of a molecule of the analyte attached to a particle located therein and exposed to a flowing aqueous medium; •a second zone in which a corresponding stream of aqueous droplets is generated from the aqueous medium and the nucleotide stream and wherein at least some of the droplets contain a single nucleotide and •a third zone in which each droplet is stored and/or interrogated to reveal a property characteristic of the single nucleotide it may contain; characterised in that the first zone comprises a microfluidic channel through which the aqueous medium flows and the location comprises a hollow seating in a wall thereof to which suction can be applied and into which the particle can be close-fitted. 2. The apparatus as claimed in claim 1 , characterised in that the hollow seating is located immediately upstream of the second zone.3. The apparatus as claimed in claim 1 , characterised in that the particle comprises a bead having a surface to which the analyte molecule can be physically or chemically bound.4. The apparatus as claimed in claim 1 , characterised in that the digestion method is selected from exonucleolysis claim 1 , phosphorolysis or pyrophosphorolysis.5. The apparatus as claimed in claim 1 , characterised in that the third zone includes a laser and a photodetector to detect Raman-scattered light.6. The apparatus as claimed in claim 1 , characterised by being capable of processing an aqueous medium which in at least one of the second or third zones contains at least one single-nucleotide probe selective for one of the nucleobase types from which the analyte is constituted; said probe(s) being capable of fluorescing substantially only after it has captured a single nucleotide and undergone subsequent exonucleolysis.7. The apparatus as claimed in claim 6 , characterised by further ...

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Номер записи: 76

12-01-2017 дата публикации

Methods and kits used in identifying microrna targets

Номер: US20170009293A1

Автор: Xiaolu Lim Ang Cambronne

Принадлежит: Oregon Health Science University

Described herein are methods and kits used to identify an endogenously expressed target mRNA of a microRNA of interest. The method involves the use of a dominant negative GW182 polypeptide that forms a stable complex with the target mRNA. The method further involves purifying the complex and identifying the target mRNA.

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Номер записи: 77

10-01-2019 дата публикации

AMPLIFICATION AND DETECTION OF NUCLEIC ACIDS

Номер: US20190009276A1

Автор: Bishop Joshua, Burns Andrew Arthur Paul, Lafleur Lisa K., Misner Matthew Jeremiah, Moore David Roger, Wheeler Maxwell

Принадлежит:

The present disclosure relates to a sample assessment device. By way of example, the sample assessment device may include a substrate including a sample application region; an amplification region comprising a plurality of amplification reagents; a waste region comprising an entrance fluidically coupled to the amplification region and extending away from the amplification region; and a detection region spaced apart from the amplification region. The sample assessment device may also include a valve coupled to the substrate and configured to separate the amplification region from the detection region in a closed configuration, wherein the amplification region and the valve are positioned on the sample assessment device between the sample application region and the detection region and wherein the sample assessment device is configured to permit lateral flow from the amplification region to the detection region when the valve is in an open configuration. 115-. (canceled)16. A sample assessment device comprising:a substrate, wherein the substrate comprises:a sample application region;an amplification region;a waste region comprising an entrance fluidically coupled to the amplification region and extending away from the amplification region; anda detection region spaced apart from the amplification region; anda valve coupled to the substrate and configured to separate the amplification region from the detection region in a closed configuration, wherein the amplification region and the valve are positioned on the sample assessment device between the sample application region and the detection region and wherein the sample assessment device is configured to permit lateral flow from the amplification region to the detection region when the valve is in an open configuration.17. The device of claim 16 , wherein the waste region extends away from an elongated strip claim 16 , wherein the sample application region claim 16 , the amplification region claim 16 , and the ...

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Номер записи: 78

27-01-2022 дата публикации

DNA stabilization of RNA

Номер: US20220025362A1

Автор: Kathleen Danenberg, Patrick Soon-Shiong, Shahrooz Rabizadeh

Принадлежит: Nantomics LLC

RNA from a biological fluid is stabilized during isolation and/or storage using DNA. In especially preferred aspects, the RNA is cfRNA and/or ctRNA, and the biological fluid is blood.

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Номер записи: 79

27-01-2022 дата публикации

Methods for non-invasive prenatal ploidy calling

Номер: US20220025456A1

Автор: Allison Ryan, Bernhard Zimmermann, George Gemelos, Johan Baner, Matthew Hill, Matthew Rabinowitz, Milena Banjevic, Styrmir Sigurjonsson, Zachary Demko

Принадлежит: Natera Inc

The present disclosure provides methods for determining the ploidy status of a chromosome in a gestating fetus from genotypic data measured from a mixed sample of DNA comprising DNA from both the mother of the fetus and from the fetus, and optionally from genotypic data from the mother and father. The ploidy state is determined by using a joint distribution model to create a plurality of expected allele distributions for different possible fetal ploidy states given the parental genotypic data, and comparing the expected allelic distributions to the pattern of measured allelic distributions measured in the mixed sample, and choosing the ploidy state whose expected allelic distribution pattern most closely matches the observed allelic distribution pattern. The mixed sample of DNA may be preferentially enriched at a plurality of polymorphic loci in a way that minimizes the allelic bias, for example using massively multiplexed targeted PCR.

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Номер записи: 80

08-01-2015 дата публикации

Method for the synthesis of a bifunctional complex

Номер: US20150011434A1

Автор: Alex Haahr Gouliaev, Anette Holtmann, Christian Klarner Sams, Eva Kampmann Olsen, Henrik Pedersen, Jakob Felding, Kim Birkebaek Jensen, Mikkel Dybro Lundorf, Per-Ola Freskgard, Sanne Birkebaek JENSEN, Soeren Nyboe Jakobsen, Thomas Franch

Принадлежит: Nuevolution AS

Disclosed is a method for obtaining a bifunctional complex comprising a display molecule part and a coding part, wherein a nascent bifunctional complex comprising a chemical reaction site and a priming site for enzymatic addition of a tag is reacted at the chemical reaction site with one or more reactants, and provided with respective tag(s) identifying the reactant(s) at the priming site is using one or more enzymes.

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Номер записи: 81

14-01-2016 дата публикации

Novel compositions, methods and kits for blood typing

Номер: US20160010153A1

Автор: Elena GRIGORENKO, Marion LAIG

Принадлежит: Life Technologies Corp

The present disclosure provides compositions, methods, and kits for typing of blood groups by their genotype. In some embodiments, the compositions, methods, and kits include one or more genotype reference nucleic acids.

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Номер записи: 82

14-01-2021 дата публикации

Labelled nucleotides

Номер: US20210009623A1

Автор: John Milton, Silke Ruediger, Xiaohai Liu, Xiaolin Wu

Принадлежит: Illumina Cambridge Ltd

The invention provides a nucleotide or nucleoside having a base attached to a detectable label via a cleavable linker, characterised in that the cleavable linker contains a moiety selected from the group comprising: Formula (I) (wherein X is selected from the group comprising O, S, NH and NQ wherein Q is a C 1-10 substituted or unsubstituted alkyl group, Y is selected from the group comprising O, S, NH and N(allyl), T is hydrogen or a C 1-10 substituted or unsubstituted alkyl group and * indicates where the moiety is connected to the remainder of the nucleotide or nucleoside).

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Номер записи: 83

11-01-2018 дата публикации

ORGANISM IDENTIFICATION PANEL

Номер: US20180010167A1

Автор: Blaschke-Bonkowsky Anne Jeannette, Poritz Mark Aaron

Принадлежит:

Methods and containers are provided for identifying a species, illustratively a bacterial species. Illustrative methods comprise amplifying various genes in the nucleic acid from the bacterial species in a single reaction mixture using pairs of outer first-stage primers designed to hybridize to generally conserved regions of the respective genes to generate a plurality of first-stage amplicons, dividing the reaction mixture into a plurality of second-stage reactions, each using a unique pair of second-stage primers, each pair of second-stage primers specific for a target bacterial species or subset of bacterial species, detecting which of the second-stage reactions amplified, and identifying the bacterial species based on second-stage amplification. Methods for determining antibiotic resistance are also provided, such methods also using first-stage primers for amplifying genes known to affect antibiotic resistance a plurality of the second-stage reactions wherein each pair of second-stage primers specific for a specific gene for conferring antibiotic resistance. 131-. (canceled)32. A method for identifying a bacterial species comprising the steps ofobtaining a sample containing the bacterial species,amplifying, in a single reaction mixture containing nucleic acid from the sample, a plurality of bacterial genes using pairs of outer first-stage primers designed to hybridize to generally conserved regions of the bacterial genes to generate a plurality of first-stage amplicons,dividing the reaction mixture into a plurality of second-stage reactions,subjecting each of a plurality of the second-stage reactions to amplification conditions, wherein each of the plurality of second-stage reactions uses a pair of second-stage primers, each pair of second-stage primers specific for a different target bacterial species or group of bacterial species,generating an amplification curve for each of the second-stage reactions that amplified,determining a crossing point for each ...

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Номер записи: 84

11-01-2018 дата публикации

CO-DETECTION AND ASSOCIATION OF MULTIPLE GENES FROM THE SAME GENOME IN A SAMPLE

Номер: US20180010170A1

Автор: Anderson Gary, Bai Jianfa, Liu Xuming

Принадлежит:

The present invention is concerned with PCR-based detection methods and kits for the identification, differentiation, and quantification of different bacterial strains (e.g., Gram-negative bacterial strains), and also association of two or more PCR-positive genes to a single genome. The methods generally comprise carrying out PCR reactions using at least a first PCR primer set and/or probe for at least one target nucleic acid; and a second PCR primer set and/or probe for at least a second target nucleic acid. Positive PCR reaction products are then detected to determine test samples containing positive PCR reaction products for both the first and second target nucleic acids. This information can be used to calculate the gene association rate to determine whether the sample contains, for example, Shiga toxin-producing of the O-type serogroup. 1. A method for co-detection of two or more target nucleic acid sequences in a sample suspected of or potentially containing Gram-negative bacteria , and determination of association of said target nucleic acid sequences as originating from a single genome or from two or more genomes , said method comprising:providing a sample suspected of or potentially containing said target nucleic acid sequences; a first PCR primer set and/or probe corresponding to a first target nucleic acid; and', 'a second PCR primer set and/or probe corresponding to a second target nucleic acid;, 'forming a reaction mixture comprising at least a portion of said sample and PCR primers and/or probes corresponding to said two or more target nucleic acid sequences, said PCR primers and/or probes comprisingdividing said reaction mixture into a plurality of individual reaction volumes;amplifying said reaction volumes to yield amplified reaction volumes comprising positive PCR reaction products; anddetermining the number of said amplified reaction volumes containing both positive PCR reaction products for said first target nucleic acid and positive PCR reaction ...

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Номер записи: 85

11-01-2018 дата публикации

METHODS AND COMPOSITIONS FOR CLONING CIRCULAR RNA

Номер: US20180010175A1

Автор: Cheng Man

Принадлежит:

Methods and compositions for in situ detection of circular RNA in a tissue sample are provided. 1. A method of in situ detection of circular RNA in a tissue or cell sample , the method comprising ,contacting the sample in situ with a strand-displacing RNA polymerase, without initially circularizing nucleic acids in the sample and without contacting circular nucleic acids to the sample, under conditions to allow for rolling circle amplification (RCA) of circular RNA in the sample, if present, to form an RCA amplicon; anddetecting the presence, absence or sequence of the RCA amplicon, thereby detecting circular RNA in the tissue sample.2. The method of claim 1 , wherein the strand displacing polymerase is primed by endogenous nucleic acid molecules in the sample.3. The method of claim 1 , further comprising add primer oligonucleotides to the sample and wherein the strand displacing polymerase is primed by primer oligonucleotides.4. The method of claim 3 , wherein the primer oligonucleotides are random or degenerate oligonucleotides.5. The method of claim 1 , wherein the strand-displacing RNA polymerase is a reverse transcriptase.6. The method of claim 1 , wherein the strand-displacing RNA polymerase is an RNA-dependent RNA polymerase.7. The method of claim 1 , further comprising contacting the sample with an exonuclease before the contacting and the detecting claim 1 , thereby degrading linear nucleic acids.8. The method of claim 1 , wherein the detecting comprises hybridizing a polynucleotide probe to the RCA amplicon.9. The method of claim 1 , wherein the hybridizing comprises fluorescence in situ hybridization (FISH).10. The method of claim 8 , wherein the polynucleotide probe is linked to a detectable label.11. The method of claim 10 , wherein the detectable label comprises a fluorophore.12. The method of claim 1 , wherein the detecting comprises nucleotide sequencing the RCA amplicon.13. The method of claim 1 , wherein the rolling circle amplification introduces ...

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Номер записи: 86

11-01-2018 дата публикации

METHODS AND SYSTEMS FOR SEQUENCING LONG NUCLEIC ACIDS

Номер: US20180010180A1

Автор: Matsuzaki Hajime, Mei Rui, Zhou Wei

Принадлежит:

The present invention provides methods and systems for sequencing long nucleic acid fragment. The present invention also provides a method of sequencing a target polynucleotide with fewer probes. Further, the present invention provides a method of sequencing a target polynucleotide with longer reads. Locus-specific, ligation-assisted sequencing/genotyping method and ligation-captured sequencing method are also provided in the present invention. The methods of the present invention allow low-cost, high-throughput and accurate sequencing of nucleic acids. 1. A method for sequencing a target nucleic acid , comprising:sequencing one or more bases of a target nucleic acid by extending a first probe hybridized to the target nucleic acid to generate a first extension product, thereby obtaining a first sequence read to determine the sequence of the target nucleic acid by performing a sequencing reaction, wherein the target nucleic acid is from a plurality of target nucleic acid fragments.2. The method of claim 1 , prior to said sequencing claim 1 , further comprising:removing a first cap from the first probe by cleaving a cleavage site at a 3′ end of the first probe, wherein the first probe comprises:(i) a capture probe from a plurality of capture probes; and 'wherein the capture probe and the first solution are ligated.', '(ii) a first solution probe from a plurality of first solution probes, each of the plurality of first solution probes comprising the first cap on the 3′ end linked via the cleavage site;'}3. The method of claim 2 , prior to said removing claim 2 , further comprising:adding a second cap to any of the plurality of capture probes not ligated to any of the plurality of first solution probes.4. The method of claim 3 , prior to said adding claim 3 , further comprising:ligating the capture probe and the first solution probe, thereby forming the first probe.5. The method of claim 4 , prior to said ligating claim 4 , further comprising:hybridizing the target ...

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Номер записи: 87

10-01-2019 дата публикации

FLUORESCENT POLYMERASE ENZYME SUBSTRATES HAVING PROTEIN SHIELDS

Номер: US20190010183A1

Автор: Bjornson Keith, BROGLEY Louis, Hanes Jeremiah, Kamtekar Satwik, Miller Erik, SEBO Lubomir

Принадлежит:

Compositions, methods, and systems are provided for fluorescent polymerase enzyme substrates comprising protein shields for improving enzyme photostability in single molecule real time sequencing. Fluorescent polymerase enzyme substrates of the invention have a protein shield between the fluorescent dye moieties and nucleotide moieties of the polymerase enzyme substrate. The polymerase enzyme substrates have a nucleotide component and a dye component, each attached to a protein. The attachments can be covalent. The protein can, for example, prevent the direct interaction of the fluorescent dye moiety with the enzyme when carrying out nucleotide synthesis, preventing photodamage to the enzyme. The polymerase enzyme substrates of the invention can have multiple dyes and multiple nucleotide moieties. 1. A polymerase enzyme substrate comprising:a protein comprising at least 60 amino acids;a nucleotide component comprising at least one nucleoside polyphosphate attached through its phosphate portion to a first position on the protein;a dye component comprising at least one fluorescent dye moiety attached to a second position on the protein.2. The polymerase enzyme substrate of wherein the substrate is connected through covalent attachments.3. The polymerase enzyme substrate of wherein the protein comprises 60 to 1 claim 1 ,000 amino acids.4. The polymerase enzyme substrate of wherein the protein comprises 80 to 600 amino acids.5. The polymerase enzyme substrate of wherein the nucleotide component and dye component are covalently attached to the protein.6. The polymerase enzyme substrate of wherein the nucleotide component comprises two or more nucleoside phosphates.7. The polymerase enzyme substrate of wherein the substrate has 2 claim 1 , 3 claim 1 , 4 claim 1 , 5. 6 claim 1 , 7 claim 1 , or 8 nucleotide phosphates.8. The polymerase enzyme substrate of wherein the dye component comprises two or more fluorescent dye moieties.9. The polymerase enzyme substrate of wherein ...

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Номер записи: 88

14-01-2021 дата публикации

Modified helicases

Номер: US20210009971A1

Автор: Andrew John Heron, Elizabeth Jayne Wallace, James White, Mark Bruce, Ruth Moysey, Szabolcs SOEROES

Принадлежит: Oxford Nanopore Technologies PLC

The invention relates to a new method of characterising a target polynucleotide. The method uses a pore and a Dda helicase. The helicase controls the movement of the target polynucleotide through the pore. The invention also relates to modified Dda helicases which can be used to control the movement of polynucleotides and are particularly useful for sequencing polynucleotides.

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Номер записи: 89

14-01-2021 дата публикации

Method for Introducing Mutations

Номер: US20210010008A1

Автор: BURKE Catherine M., DARLING Aaron E., IMELFORT Michael, MONAHAN Leigh G., To Joyce

Принадлежит:

The present invention relates to a method for introducing mutations into at least one target nucleic acid molecule comprising (a) providing at least one sample comprising at least one target nucleic acid molecule; and (b) amplifying the at least one target nucleic acid molecule using a low bias DNA polymerase. The present further relates to a use of a low bias DNA polymerase in a method for introducing mutations into one or more nucleic acid molecule(s), a group of sample tags, a method for designing the group of sample tags, a computer readable medium, and a method for preferentially amplifying target nucleic acid molecules. 1. A method for introducing mutations into at least one target nucleic acid molecule comprising:a. providing at least one sample comprising at least one target nucleic acid molecule; andb. amplifying the at least one target nucleic acid molecule using a low bias DNA polymerase.2. Use of a low bias DNA polymerase in a method for introducing mutations into at least one target nucleic acid molecule.3. The use of claim 2 , wherein the method for introducing mutations into at least one target nucleic acid molecule comprises:a. providing at least one sample comprising at least one target nucleic acid molecule; andb. amplifying the at least one target nucleic acid molecule using a low bias DNA polymerase.4. The method or use of any one of the preceding claims claim 2 , wherein the mutations are substitution mutations.5. The method or use of any one of the preceding claims claim 2 , wherein the low bias DNA polymerase mutates adenine claim 2 , thymine claim 2 , guanine claim 2 , and cytosine nucleotides in the at least one target nucleic acid molecule at a rate ratio of 0.5-1.5:0.5-1.5:0.5-1.5:0.5-1.5 claim 2 , 0.6-1.4:0.6-1.4:0.6-1.4:0.6-1.4 claim 2 , 0.7-1.3:0.7-1.3:0.7-1.3:0.7-1.3 claim 2 , 0.8-1.2:0.8-1.2:0.8-1.2:0.8-1.2 claim 2 , or around 1:1:1:1 respectively.6. The method or use of any one of the preceding claims claim 2 , wherein the low bias ...

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Номер записи: 90

14-01-2021 дата публикации

METHOD AND APPARATUS TO NORMALIZE QUANTITATIVE READOUTS IN SINGLE-CELL EXPERIMENTS

Номер: US20210010061A1

Автор: Mendez Pedro, Ruff David

Принадлежит:

Provided herein are methods and systems for detection of nucleic acids for single cell samples. As part of the detection, a unique step of normalization of different single cell samples is included. One embodiment of the method includes i) selecting one or more target nucleic acid sequence of interest in an individual cell, where the target nucleic acid sequence is complementary to a nucleic acid in a cell; ii) providing a sample having a plurality of individual single cells and encapsulating one or more individual cell(s); iii) providing a sample normalization component to one or more encapsulated cell, where the normalization component comprises an exogenous nucleic acid having a known sequence; iv) providing nucleic acid primers for the target nucleic acid and the exogenous nucleic acid; v) providing a protease to each encapsulated cell and incubating the encapsulated cell with the protease in the drop to produce a cell lysate; vi) performing a nucleic acid amplification reaction to form an amplification product from the nucleic acid of a single cell, where the amplification product comprise amplicons of one or more target nucleic acid sequence and an amplicon for the exogenous nucleic acid; and vii) comparing the amplification products from the target amplicons and the exogenous nucleic acid amplicons and determining the copy number or sequence of the target nucleic acid in a single cell. 1. A method for nucleic acid detection for single cell samples , the method comprising , independent of order presented , the following steps:i) selecting one or more target nucleic acid sequence of interest in an individual cell, wherein the target nucleic acid sequence is complementary to a nucleic acid in a cell;ii) providing a sample having a plurality of individual single cells and encapsulating one or more individual cell(s);iii) providing a sample normalization component to one or more encapsulated cell, wherein the normalization component comprises an exogenous nucleic ...

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Номер записи: 91

14-01-2021 дата публикации

ENRICHMENT OF NUCLEIC ACIDS

Номер: US20210010064A1

Автор: Bau Haim H., Liu Changchun, Mauk Michael G., SONG Jinzhao, Van Der Oost John, WIETZE HEGGE Jorrit

Принадлежит:

Provided are methods directed to enriching nucleic acids in a biological sample. These methods, in some embodiments can discriminately enrich the abundance of low-copy nucleic acids relative to higher-copy nucleic acids. In some embodiments, the methods provided can enrich a low-copy number mutant allele associated with a disease state, thus allowing early detection and optimized treatment. In other embodiments, the methods can be used for detection of particular molecules, such as antigens, in a sample. 1. A method of enriching a target nucleic acid in a sample , comprising:a. contacting the sample with a guide nucleic acid having a sufficiently complementary sequence to a non-target nucleic acid to allow hybridization of the guide nucleic acid and the non-target nucleic acid to form a guide/non-target hybrid;b. contacting the sample with an endonuclease having an affinity for the guide/non-target hybrid; andc. amplifying the target nucleic acid or incubating the sample.2. (canceled)3. (canceled)4. The method of claim 1 , wherein the target nucleic acid is a low-copy nucleic acid.5. The method of claim 1 , wherein the target nucleic acid is less than about 10% as abundant as the non-target nucleic acid.6. The method of claim 1 , wherein the target nucleic acid is less than about 0.1% as abundant as the non-target nucleic acid.7. The method of claim 1 , further comprising removing the endonuclease before amplifying the target nucleic acid.8. (canceled)9. The method of claim 1 , wherein amplifying the target nucleic acid comprises polymerase chain reaction (PCR) claim 1 , digital drop PCR claim 1 , loop-mediated isothermal amplification (LAMP) claim 1 , recombinase polymerase amplification (RPA) claim 1 , or any combination thereof.10. The method of claim 1 , wherein the target nucleic acid comprises a mutation or is a variant associated with a disease.11. The method of claim 1 , wherein the target nucleic acid and the non-target nucleic acid are from different ...

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Номер записи: 92

14-01-2021 дата публикации

METHODS AND REAGENTS FOR ENRICHMENT OF NUCLEIC ACID MATERIAL FOR SEQUENCING APPLICATIONS AND OTHER NUCLEIC ACID MATERIAL INTERROGATIONS

Номер: US20210010065A1

Автор: Li Tan, SALK Jesse J., WILLIAMS Lindsey Nicole

Принадлежит:

The present technology relates generally to methods and compositions for targeted nucleic acid sequence enrichment, as well as uses of such enrichment for error-corrected nucleic acid sequencing applications and other nucleic acid sequence interrogations. In some embodiments, provided methods provide non-amplification based targeted enrichment strategies compatible with the use of molecular barcodes for error correction. Other embodiments provide methods for non-amplification based targeted enrichment strategies compatible with direct digital sequencing (DDS) and other sequencing strategies (e.g., single molecule sequencing modalities and interrogations) that do not use molecular barcoding. 1. A method for enriching target nucleic acid material , comprising:providing a nucleic acid material;cutting the nucleic acid material with one or more targeted endonucleases so that a target region of predetermined length is separated from the rest of the nucleic acid material;enzymatically destroying non-targeted nucleic acid material;releasing the target region of predetermined length from the targeted endonuclease; andanalyzing the cut target region.2. The method of claim 1 , wherein enzymatically destroying non-targeted nucleic acid material comprises providing an exonuclease enzyme.3. The method of claim 1 , wherein enzymatically destroying non-targeted nucleic acid material comprises providing one or more of an exonuclease enzyme and an endonuclease enzyme.4. The method of claim 1 , wherein the destroying comprises at least one of enzymatic digestion and enzymatic cleavage.5. The method of any one of - claim 1 , wherein the one or more targeted endonucleases remain bound to the target region during the enzymatically destroying step.6. The method of any one of - claim 1 , wherein at least one targeted endonuclease is a ribonucleoprotein complex comprising a capture label claim 1 , and wherein the target region of predetermined length is physically separated from the rest ...

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Номер записи: 93

14-01-2021 дата публикации

PREPARATION OF NUCLEIC ACID LIBRARIES FROM RNA AND DNA

Номер: US20210010073A1

Автор: Aravanis Alex, Cao Dan, Xu Hongxia

Принадлежит:

Some embodiments of the methods and compositions provided herein relate to the preparation and use of nucleic acid libraries derived from RNA and DNA. In some embodiments, a nucleic acid library can be prepared by tagging polynucleotides derived from RNA. Some embodiments include the analysis of sequence data from such libraries. 143.-. (canceled)44. A method for preparing a library of nucleic acids comprising:(a) hybridizing a plurality of polynucleotides with a plurality of primers, wherein the plurality of polynucleotides comprises RNA and DNA;(b) extending the hybridized primers with a reverse transcriptase; and(c) generating a library of nucleic acids from the extended primers and the DNA.45. The method of claim 44 , further comprising (d) sequencing the library of nucleic acids.46. The method of claim 44 , wherein the plurality of primers comprise tags.47. The method of claim 46 , further comprising (e) identifying polynucleotide sequences comprising the tags claim 46 , thereby identifying sequences derived from the RNA polynucleotides of the plurality of polynucleotides.48. The method of claim 47 , further comprising identifying polynucleotide sequences lacking the tags claim 47 , thereby identifying sequences derived from the DNA polynucleotides of the plurality of polynucleotides.49. The method of claim 44 , wherein the plurality of primers comprises different sequences.50. The method of claim 44 , wherein the plurality of primers comprises greater than 10 claim 44 ,000 different sequences.51. The method of claim 46 , wherein the plurality of primers comprises the same tag.52. The method of claim 44 , wherein the reverse transcriptase lacks a DNA-dependent polymerase activity.53. The method of claim 44 , wherein (b) is performed in the presence of the DNA polynucleotides.54. The method of claim 44 , wherein (b) comprises generating double-stranded cDNA from the extended primers.55. The method of claim 44 , wherein the plurality of polynucleotides is cell- ...

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Номер записи: 94

14-01-2021 дата публикации

RNA SEQUENCING METHODS

Номер: US20210010075A1

Автор: Almogy Gilad, OBERSTRASS Florian

Принадлежит:

Described herein are methods for determining a sequence of a region of interest from an mRNA molecule. Sequenced polynucleotides can include a barcode region, a homopolymer region (e.g., a poly-A region), and a target region associated with the mRNA molecule. According to some methods, the barcode region omits the same base present in the homopolymer region. According to some methods, extension of the primer used for sequencing is stalled within the homopolymer region. According to some methods, sequencing flow cycles and the different barcode regions of the polynucleotides configured are such that the primer is extended to the end of the barcode region across the plurality of polynucleotides before being extended into the homopolymer region. According to some methods, two primers or a cleavable primer is used to separately sequence the barcode region and the target region. 1. A method of determining a sequence of a region of interest from an mRNA molecule , comprising:hybridizing a plurality of polynucleotides derived from mRNA with a primer to form a plurality of hybridized templates, the polynucleotides comprising a barcode region, a homopolymer region comprising a plurality of contiguous and identical bases, and a target region comprising a sequence associated with a region of interest from an mRNA molecule, wherein the barcode region omits the same base present in the homopolymer region;determining the sequence of the barcode region using labeled nucleotides that lack a complement base of the same base present in the homopolymer region;extending the primer within the homopolymer region using nucleotides complementary to the base present in the homopolymer region; anddetermining the sequence of the target region using labeled nucleotides.2. The method of claim 1 , wherein the labeled nucleotides used when determining the sequence of the barcode region comprise non-terminating labeled nucleotides.3. The method of claim 1 , wherein the nucleotides used when ...

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Номер записи: 95

14-01-2016 дата публикации

Nanopore sequencing methods

Номер: US20160011169A1

Автор: Jonas Korlach, Stephen Turner

Принадлежит: Pacific Biosciences of California Inc

Methods are provided for sequencing of nucleic acid templates using nanopores. The rate of transport of the template nucleic acids through the nanopore is controlled using a polymerase enzyme having two slow kinetic steps. Methods are provided for sequencing hemi-natural nucleic acids such as hemi-genomic DNA, having two complementary strands, one a natural sequence and the other a synthetic sequence. The identification of modified bases can be enhanced by comparing the sequencing information from the natural sequence, which has, for example, natural base modifications, with the synthetic sequence, which typically has no base modifications. The presence and identity of a modified base can be determined by monitoring kinetics, for example the kinetics of polymer meditated nucleic acid synthesis.

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Номер записи: 96

14-01-2016 дата публикации

Method and system for substance detection with a magnetic sensor

Номер: US20160011182A1

Автор: Jiaoming Qiu

Принадлежит: Zepto Life Technology Inc

Methods, systems and programing for substance detection with a magnetic sensor are presented. In one example, a magnetic sensor having one or more layers is formed on a base for sensing a magnetic field created by magnetic particles present in proximity to the magnetic sensor. A first end of each of a first set of strands is immobilized with respect to the magnetic sensor. A magnetic particle is attached to a second end of each of the first set of strands so that when a material containing a substance is in contact with the base, the substance causes at least some of the first set of strands to break resulting in that the magnetic particle attached to the second end of each of the at least some of the first set of strands is no longer in proximity to the magnetic sensor.

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Номер записи: 97

10-01-2019 дата публикации

NUCLEIC ACID AMPLIFICATION AND DETECTION ASSAYS

Номер: US20190010539A1

Автор: CERDA-MOYA Gustavo Andrés

Принадлежит: Cambridge Molecular Diagnostics Ltd

The present invention relates to a method and kit for amplifying and detecting a quantity of nucleic acid. The invention is particularly relevant to isothermal amplification techniques carried out on a flow based assay device. The amplified nucleic acid may be detected on the device using an optical read-out. 1. A test strip device for detecting the presence of a target analyte in a fluid applied to the device , the device comprising:i) a displacement area having a first immobilised marker which can be displaced by the presence of the target analyte to produce a first released marker, wherein the first released marker comprises a protease;ii) a signal amplification area having further immobilised markers or detectable markers which can be released by the presence of the protease;iii) optionally one or more further signal amplification areas having further immobilised markers which can be released by the presence of a released marker, wherein one or more of the signal amplification areas releases a detectable marker; andiv) one or more detection areas which can identify the presence of the detectable marker;wherein the displacement area, signal amplification area(s) and detection area(s) are connected on a porous material such that fluid can flow from the displacement area through the signal amplification area(s) and into the detection area(s) upon application of the fluid to the device.2. The device according to wherein the target analyte is a nucleic acid.3. The device according to wherein the first immobilised marker is a partly double stranded nucleic acid.4. The device according to wherein the device contains three or more signal amplification areas.5. (canceled)6. (canceled)7. The device according to wherein the detectable marker contains an enzyme.8. The device according to wherein the detectable marker carries a detectable label.9. The device according to wherein the detection area performs a colourimetric assay.10. The device according to wherein the ...

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Номер записи: 98

10-01-2019 дата публикации

HIGH THROUGHPUT GENOME SEQUENCING ON DNA ARRAYS

Номер: US20190010542A1

Автор: Callow Matthew J., Drmanac Radoje, Drmanac Snezana

Принадлежит: Complete Genomics Inc.

The present invention is directed to methods and compositions for acquiring nucleotide sequence information of target sequences using adaptors interspersed in target polynucleotides. The sequence information can be new, e.g. sequencing unknown nucleic acids, re-sequencing, or genotyping. The invention preferably includes methods for inserting a plurality of adaptors at spaced locations within a target polynucleotide or a fragment of a polynucleotide. Such adaptors may serve as platforms for interrogating adjacent sequences using various sequencing chemistries, such as those that identify nucleotides by primer extension, probe ligation, and the like. Encompassed in the invention are methods and compositions for the insertion of known adaptor sequences into target sequences, such that there is an interruption of contiguous target sequence with the adaptors. By sequencing both “upstream” and “downstream” of the adaptors, identification of entire target sequences may be accomplished. 1. (canceled)2. A method of preparing an array of fragments of a target nucleic acid to be sequenced , the method comprising:(a) providing a plurality of double-stranded DNA molecules each containing a target sequence from the target nucleic acid;(b) introducing a nick into one of the strands in each of the double-stranded DNA molecules at an initial site, exposing a free 5′ strand and a free 3′ strand at the site;(c) treating the nicked DNA molecules with a DNA polymerase so as to extend the free 3′ strand and displace or degrade the free 5′ strand, thereby moving the nick along the nicked strand to a new position downstream from the initial site;(d) disabling the DNA polymerase;(e) cleaving the DNA molecules at the new position, thereby forming a gap in both strands of the DNA molecules;(f) inserting a downstream oligonucleotide adaptor into the gap of each DNA molecule to form a nucleic acid construct; and(g) arraying the constructs on a solid surface for nucleic acid sequencing.3. The ...

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Номер записи: 99

10-01-2019 дата публикации

METHODS FOR SIMULTANEOUS AMPLIFICATION OF TARGET LOCI

Номер: US20190010543A1

Автор: BABIARZ Joshua, CONSTANTIN Tudor Pompiliu, EUBANK Lane A., Gemelos George, HILL Matthew Micah, KIRKIZLAR Huseyin Eser, Rabinowitz Matthew, SAKARYA Onur, SIGURJONSSON Styrmir, Zimmermann Bernhard

Принадлежит: Natera, Inc.

The invention provides methods for simultaneously amplifying multiple nucleic acid regions of interest in one reaction volume as well as methods for selecting a library of primers for use in such amplification methods. The invention also provides library of primers with desirable characteristics, such as minimal formation of amplified primer dimers or other non-target amplicons. 1. A method for determining a transplant status , comprising(a) obtaining a nucleic acid sample from a blood, plasma, or serum sample of a transplant recipient, wherein the nucleic acid sample comprises a mixture of cell-free DNA from a transplant and cell-free DNA from the transplant recipient;(b) amplifying at least 100 different target loci from the nucleic acid sample by contacting the nucleic acid sample with a library of at least 100 non-immobilized, non-identical primers that hybridize to the at least 100 different target loci to produce a reaction mixture, and subjecting the reaction mixture to PCR conditions to produce amplification products comprising target amplicons, wherein at least 50% of the amplification products each comprises at least one of the target loci;(c) measuring an absolute or relative amount of transplant cell-free DNA from the amplification products, and determining a transplant status based on the measured amount of transplant cell-free DNA, wherein the transplant status is transplant rejection, tolerance, non-rejection based allograft injury, transplant function, transplant survival, chronic transplant injury, or tittering of pharmacological immunosuppression.2. The method of claim 1 , wherein the length of the target amplicons is less than 100 nucleotides.3. The method of claim 1 , wherein a range of melting temperatures among the different primers in the library is less than 10° C.4. The method of claim 1 , wherein a range of melting temperatures among the different primers in the library is less than 5° C.5. The method of claim 1 , further comprising ...

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Номер записи: 100