Abstract: A method for identification of the most abundant oligonucleotide species in a library of oligonucleotides comprising more than 106 oligonucleotide species, wherein the oligonucleotides of the library of oligonucleotides are not inherently capable of exponential amplification by PCR and include a coding sequence, which is characteristic of one oligonucleotide specie of the library of oligonucleotides and only one fixed sequence, which is present in a plurality of oligonucleotide species of the library of oligonucleotides, said fixed sequence being located on a 5? side of the coding sequence, and wherein the method comprises specifically amplifying the sequence of a hybridised oligonucleotide species using the steps of: incubating the library of oligonucleotides under conditions of hybridization such as to allow complementary coding sequences to hybridize and form hybridized oligonucleotide species; extending a 3? end of one or more of the hybridised oligonucleotide species, having only one fixed sequence, to p
Abstract: Described herein are methods and kits for detecting the presence or absence of gene dysregulations such as those arising from gene fusions and/or chromosomal abnormalities, e.g. translocations, insertions, inversions and deletions. The methods, compositions and kits are useful for detecting mutations that cause the differential expression of a 5? portion of a target gene relative to the 3? region of the target gene. The average expression of the 5? portion of the target gene is compared with the average expression of the 3? portion of the target gene to determine an intragenic differential expression (IDE). The IDE can then be used to determine if a dysregulation or a particular disease (or susceptibility to a disease) is present or absent in a subject or sample.
Abstract: A method of sample analysis is provided. In certain embodiments, the method involves: a) amplifying a product from a sample that comprises both wild type copies of a genomic locus and mutant copies of the genomic locus that have a point mutation relative to said wild type copies of the genomic locus, to produce an amplified sample, where: i. the amplifying is done using a first primer and a second primer; and ii. the first primer comprises a 3? terminal nucleotide that base pairs with the point mutation and also comprises a nucleotide sequence that is fully complementary to a sequence in the locus with the exception of a single base mismatch within 6 bases of the 3? terminal nucleotide; and b) detecting the presence of said product in said amplified sample using a flap assay that employs an invasive oligonucleotide. A kit for performing the method is also provided.
Type:
Grant
Filed:
September 12, 2019
Date of Patent:
August 17, 2021
Assignee:
EXACT SCIENCES DEVELOPMENT COMPANY, LLC
Inventors:
Hongzhi Zou, Graham P. Lidgard, Michael J. Domanico, Hatim Allawi
Abstract: The present invention concerns preparation of DNA molecules, such as a library, using a stem-loop oligonucleotide. In particular embodiments, the invention employs a single reaction mixture and conditions. In particular, at least part of the inverted palindrome is removed during the preparation of the molecules to facilitate amplification of the molecules. Thus, in specific embodiments, the DNA molecules are suitable for amplification and are not hindered by the presence of the palindrome.
Type:
Grant
Filed:
December 21, 2018
Date of Patent:
July 27, 2021
Assignee:
Takara Bio USA, Inc.
Inventors:
Vladimir L. Makarov, Emmanuel Kamberov, Brendan J. Tarrier
Abstract: Provided herein are methods and compositions for single cell characterization using affinity-oligonucleotide conjugates. Provided herein are methods and compositions for single cell charaterization using tetramer-oligonucleotide conjugates.
Type:
Grant
Filed:
August 3, 2019
Date of Patent:
July 13, 2021
Assignee:
AbVitro LLC
Inventors:
Francois Vigneault, Adrian Wrangham Briggs, Stephen J. Goldfless, Brian J. Belmont
Abstract: A method for identifying the genetic relatedness of Liriodendron hybrids is provided, including: 1) extracting DNA of Liriodendron hybrid leaves; 2) performing PCR detection; and 3) identifying the genetic relatedness of Liriodendron spp. according to the size of PCR products amplified with different primers. A special kit for the method for identifying the genetic relatedness of Liriodendron hybrids is further provided, including a primer pair 18.2 and a primer pair 700. Moreover, an application of a special kit for identification of the genetic relatedness of Liriodendron hybrids is provided.
Abstract: Methods and devices are provided for simultaneously amplifying a plurality of sample wells for a predetermined amount of amplification, detecting whether amplification has occurred in a first set of the wells, amplifying for an additional amount of amplification and detecting whether amplification has occurred in a second set of the wells. Methods are also provided for analyzing a target nucleic acid sequence using melt curves that were generated in a plurality of amplification cycles.
Type:
Grant
Filed:
July 17, 2018
Date of Patent:
June 22, 2021
Assignee:
BioFire Diagnostics, LLC
Inventors:
Randy P. Rasmussen, Robert John Crisp, Andrew Clinton Hemmert, Elizabeth Barker Campbell, Thomas Charles Robbins, David J. Eyre
Abstract: A method of maintaining contiguity in chromosomal DNA following treatment with a tagmentase. Conditions are selected such that the tagmentase does not release from the DNA, and thus forms a bridge linking DNA segments that have the same relationship (haplotype) as occurred in the genomic DNA. Thus the tagmentase step can occur in bulk (before partitions are formed). The resulting tagmentase-bridged DNA segments can be added to partitions maintaining the bridged segments until they are introduced into different partitions. Once in partitions, the contiguous DNA segments can be barcoded with a partition-specific barcode, thereby allowing for later identification of contiguous DNA after sequencing in bulk (after partitions contents are merged).
Abstract: Described herein are methods, compositions and kits directed to the detection of gene dysregulations such as those arising from gene fusions and/or chromosomal abnormalities, e.g., translocations, insertions, inversions and deletions. Samples containing dysregulated gene(s) of interest may show independent expression patterns for the 5? and 3? regions of the gene. The methods, compositions and kits are useful for detecting mutations that cause the differential expression of a 5? portion of a target gene relative to the 3? region of the target gene.
Type:
Grant
Filed:
December 9, 2019
Date of Patent:
June 1, 2021
Assignee:
Quest Diagnostics Investments LLC
Inventors:
Heather R. Sanders, Maher Albitar, Aurelia Meloni-Ehrig
Abstract: The present disclosure relates to systems and methods for purifying nucleic acid. In particular, the present disclosure relates to systems and methods for purifying nucleic acids using metal or metal oxide compositions.
Abstract: The disclosed embodiments generally relate to a method and system to synthesize a target molecule within a droplet. In an exemplary embodiment, a first microfluidic device configured to contact a polynucleotide-containing component from a sample with lysis reagents to form a first droplet. The lysis reagents include an enzyme having protease activity. The first droplet is encapsulated with an immiscible carrier fluid. A collection reservoir is provided to receive and incubate the first droplet for a first duration of time. The first duration of time is sufficient to inactivate the enzyme of the lysis reagent. A second microfluidic device is provided to receive the first droplet and add nucleic acid synthesis reagent to thereby form a second nucleic acid synthesis droplet in the immiscible carrier fluid. Finally, a reaction chamber is provided to synthesize the target polynucleotide within the second nucleic acid synthesis droplet.
Type:
Grant
Filed:
April 11, 2019
Date of Patent:
May 11, 2021
Assignee:
The Regents of the University of California
Inventors:
Adam R. Abate, Dennis Jay Eastburn, Adam R. Sciambi
Abstract: The present disclosure provides a system and method for the detection of rare mutations and copy number variations in cell free polynucleotides. Generally, the systems and methods comprise sample preparation, or the extraction and isolation of cell free polynucleotide sequences from a bodily fluid; subsequent sequencing of cell free polynucleotides by techniques known in the art; and application of bioinformatics tools to detect rare mutations and copy number variations as compared to a reference. The systems and methods also may contain a database or collection of different rare mutations or copy number variation profiles of different diseases, to be used as additional references in aiding detection of rare mutations, copy number variation profiling or general genetic profiling of a disease.
Abstract: Methods of detecting aneuploidy in in vitro fertilized embryos are provided. A unique set of STR markers that can be rapidly and accurately quantified by multiplex PCR at the single cell level are used to analyze and select euploid embryos in an in vitro fertilization (IVF) setting. The markers include D13S284, D13S141, D18S54, D18S70, D21S266, D21S1951 and AMXY and are used to detect abnormalities in chromosomes 13, 18, 21 and the XY chromosomes.
Abstract: The present invention provides a low-frequency mutation enrichment sequencing method for free target DNA in plasma, comprising plasma DNA extraction and library construction, general library TT COLD PCR amplification enrichment, probe enrichment capture, PCR and sequencing of captured products, and positive and negatice double-strand error-correction low-frequency information analysis.
Type:
Grant
Filed:
February 18, 2016
Date of Patent:
May 11, 2021
Assignee:
Geneplus—Beijing
Inventors:
Xiaoxing Lv, Xin Yi, Meiru Zhao, Yanfang Guan, Tao Liu, Ling Yang
Abstract: The present disclosure provides a system and method for the detection of rare mutations and copy number variations in cell free polynucleotides. Generally, the systems and methods comprise sample preparation, or the extraction and isolation of cell free polynucleotide sequences from a bodily fluid; subsequent sequencing of cell free polynucleotides by techniques known in the art; and application of bioinformatics tools to detect rare mutations and copy number variations as compared to a reference. The systems and methods also may contain a database or collection of different rare mutations or copy number variation profiles of different diseases, to be used as additional references in aiding detection of rare mutations, copy number variation profiling or general genetic profiling of a disease.
Abstract: A PCR reaction vessel includes: a substrate; a channel formed on the substrate; a pair of filters, a first filter and a second filter, provided at respective ends of the channel; a pair of air communication ports, a first air communication port and a second air communication port, that communicate with the channel through the first filter and the second filter; a thermal cycle region formed between the first filter and the second filter in the channel; a branch point formed between the first filter and the second filter in the channel; a branched channel whose one end is connected to the branch point; and a sample introduction port formed at the other end of the branched channel.
Type:
Grant
Filed:
May 31, 2018
Date of Patent:
April 27, 2021
Assignees:
Nippon Sheet Glass Company, Limited, NATIONAL INSTITUTE OF ADVANCED INDUSTRIAL SCIENCE AND TECHNOLOGY, Go!Foton, Inc.
Abstract: Methods are provided herein for identifying rare and/or unknown DNA sequences by next-generation sequencing approaches. Isolated double-stranded (ds), single-stranded (ss), or ds/ss DNA is fragmented and the fragments are polished, phosphorylated, and tailed, as necessary. Fragmentation can be enzymatic or mechanical. A universal adapter sequence is ligated to each fragment, wherein the adapter can have a top strand without a 5? phosphate, a 3? with an —H in place of the —OH, and/or a 3? extra base complementary to any base added to the polished fragments. The ligatamers may then serve as templates for amplification using a forward primer complementary to the adapter sequence and a reverse primer targeted to the fragment sequence. Compositions produced by these methods and kits adapted for performing these methods are also described herein.
Abstract: The present application is directed to biosensors and methods for detecting a microorganism target in a sample using a mechanically interlocked nucleic acid catanane, wherein an enzyme from the microorganism target or that is activated by a molecule from the microorganism target cleaves a linkage in a first single-stranded nucleic acid ring of the catanane structure, allowing rolling-circle amplification to occur and the presence of rolling-circle amplification products indicates the presence of the microorganism in the sample.
Abstract: The disclosure provides methods for isolating nucleic acids from a biological fluid. In one aspect, the disclosure provides a method for isolating RNA. In another aspect, the disclosure provides a method for isolating DNA. In one aspect, the methods described herein utilize a protocol that combines a detergent-based initial denaturation, protease digestion, and organic extraction followed by column purification that maximizes RNA/DNA yield and preserves RNA/DNA integrity. In yet another aspect, the disclosure provides a kit for isolating RNA and/or DNA.
Type:
Grant
Filed:
December 31, 2015
Date of Patent:
April 6, 2021
Assignee:
THE ROCKEFELLER UNIVERSITY
Inventors:
Klaas Max, Karl Bertram, Kemal Akat, Thomas H. Tuschl, Jenny Li, Kimberly Bogardus