The present disclosure relates to compositions of matter and assay methods used to detect one or more target nucleic acids of interest in a sample. The compositions and methods provide signal boost upon detection of target nucleic acids of interest in less than one minute and in some instances instantaneously at ambient temperatures down to 16° C. or less, without amplification of the target nucleic acids yet allowing for massive multiplexing, high accuracy and minimal non-specific signal generation.
Presented are assay modules for performing multiplex assays that detect many influenza virus-related nucleic acids simultaneously, including those from known influenza viruses and influenza virus subtypes as well as previously unknown, emerging influenza viruses and influenza subtypes, as well as other viruses and bacteria known to cause respiratory symptoms in humans and other animals. Neither sample preparation methods nor the assays necessarily require amplification of the sample nucleic acids yet the assays retain sensitivity, allow for multiplexing, and provide low cost, minimum automated workflow and results in less than thirty minutes.
C12Q 1/70 - Measuring or testing processes involving enzymes, nucleic acids or microorganismsCompositions thereforProcesses of preparing such compositions involving virus or bacteriophage
C12Q 1/6818 - Hybridisation assays characterised by the detection means involving interaction of two or more labels, e.g. resonant energy transfer
3.
RAPID DETECTION OF KNOWN AND EMERGING INFLUENZA VIRUSES
Presented are assay modules for performing multiplex assays that detect many influenza virus-related nucleic acids simultaneously, including those from known influenza viruses and influenza virus subtypes as well as previously unknown, emerging influenza viruses and influenza subtypes, as well as other viruses and bacteria known to cause respiratory symptoms in humans and other animals. Neither sample preparation methods nor the assays necessarily require amplification of the sample nucleic acids yet the assays retain sensitivity, allow for multiplexing, and provide low cost, minimum automated workflow and results in less than thirty minutes.
G01N 33/569 - ImmunoassayBiospecific binding assayMaterials therefor for microorganisms, e.g. protozoa, bacteria, viruses
B01L 3/00 - Containers or dishes for laboratory use, e.g. laboratory glasswareDroppers
C12Q 1/70 - Measuring or testing processes involving enzymes, nucleic acids or microorganismsCompositions thereforProcesses of preparing such compositions involving virus or bacteriophage
G01N 33/58 - Chemical analysis of biological material, e.g. blood, urineTesting involving biospecific ligand binding methodsImmunological testing involving labelled substances
C07K 16/10 - Immunoglobulins, e.g. monoclonal or polyclonal antibodies against material from viruses from RNA viruses
G01N 21/69 - Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light electrically excited, e.g. electroluminescence specially adapted for fluids
4.
SYSTEMS USING CAS12 CRISPR NUCLEASES FOR SEQUENCE-SPECIFIC, DNA-INDEPENDENT DETECTION OF SSRNA
The present disclosure relates to one-RNP or two-RNP systems and methods used to detect one to many to a massively multiplexed number of target nucleic acids of interest, including ssDNA, dsDNA and ssRNA, where the detection of ssRNA is independent of the presence of DNA. The RNP in the one-RNP system and the first RNP in the two-RNP system comprise a Casl2 nuclease and a guide RNA.
The present disclosure relates to multiplex assay methods used to detect target nucleic acids of interest from several to many sources in a sample without amplification of the target nucleic acids of interest. The methods involve partitioning of first ribonucleoprotein complexes to detect short sequences from several to many loci scattered throughout a single source genome, and the addition of signal boosting second ribonucleoprotein complexes and reporter moieties.
The present disclosure relates to compositions of matter and assay methods used to detect one or more non-nucleic acid targets of interest in a sample. The compositions and methods provide signal boost upon detection of non-nucleic acid targets of interest in less than one minute and in some instances instantaneously at ambient temperatures down to 25° C. or less, allow for massive multiplexing, high accuracy, minimal non-specific signal generation, and are easily reprogrammable.
The present disclosure relates to variant engineered nucleic acid-guided nucleases that may be used in CRISPR-based cascade assay systems to detect one or more target nucleic acids in a sample. The variant nucleases comprise an activity such that double- stranded DNA substrates do not bind to or are not cleaved by variant LbCas12a nuclease, or bind to or are cleaved very slowly by the variant nuclease, however single- stranded DNA substrates can bind and are cleaved by the variant nuclease, and wherein the variant nuclease exhibits both cis- and trans-cleavage activity
The present disclosure relates to compositions of matter and assay methods used to detect one or more target nucleic acids of interest in a sample. The compositions and methods allow one to control reaction kinetics of the cascade assay by two orders of magnitude via molecular design of one of the reaction components; further, varying molecular design also allows for quantification of target nucleic acids of interest over a large range of concentrations or discriminating between extremely low copy numbers of target nucleic acids of interest.
The present disclosure relates to assay modules on which multiplex nucleic acid detection assays are performed to detect target nucleic acids of interest from several to many sources in a sample without amplification of the target nucleic acids of interest. The assay modules introduce a sample simultaneously to several to many different target-specific modalities.
The present disclosure relates to compositions of matter and methods used to activate effector nucleic acids and effector targets in vivo via a CRISPR-based cascade system. The compositions and methods achieve non-specific delivery of cascade system components to cells yet the cascade system works in a cell-specific manner.
Automated sample prep devices and methods for the use thereof is presented. The automated sample prep devices are unique in that they accomplish cell lysis, nucleic acid fragmentation, and, in some embodiments, nucleic acid purification via bead beating using two diametrically magnetized magnets. The two magnets are set apart so that the magnetic fields of the magnets interact both statically and dynamically. A sample prep chamber is positioned between the magnets in the middle of the interacting magnetic fields. The sample chamber comprises a sample, as well as lysis/binding beads and a number of metal spheres. The spheres trigger the interaction or movement between both the spheres themselves and the lysis/binding beads as the magnetic fields are altered.
The present disclosure relates to multiplex assay methods and systems used to detect several to many to a massively multiplexed number of target nucleic acids of interest in a sample without amplification of the target nucleic acids of interest. The method employs microfluidic droplet systems where each droplet is a "mini-reactor." In some embodiments, a "bulk format" configuration is used, in other embodiments, a "sequential format" configuration is used.
The present disclosure relates to multiplex assay methods and systems used to detect several to many to a massively multiplexed number of target nucleic acids of interest in a sample without amplification of the target nucleic acids of interest. The method employs microfluidic droplet systems where each droplet is a “mini-reactor.” In some embodiments, a “bulk format” configuration is used, in other embodiments, a “sequential format” configuration is used.
C12Q 1/689 - Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes for detection or identification of organisms for bacteria
The present disclosure relates to compositions of matter and assay methods used to screen for molecular dimerization events using a two-ribonucleoprotein complex signal boost assay. The compositions and methods provide a readout upon detection of dimerization of molecules and may be implemented in a high throughput manner using libraries of tens to hundreds to thousands of putative binding partners.
The present disclosure relates to stabilization variant engineered nucleic acid-guided nucleases that are used in CRISPR-based cascade assay systems to detect one or more target nucleic acids in a sample. The cascade assay systems provide signal boost upon detection of target nucleic acids without boost of the target nucleic acids. The stabilization variant engineered nucleic acid-guided nucleases are particularly useful in point-of-care applications, in the field, and as components of assay kits.
The present disclosure relates to compositions of matter and assay methods used to detect one or more non-nucleic acid targets of interest in a sample. The compositions and methods provide signal boost upon detection of non-nucleic acid targets of interest in less than one minute and in some instances instantaneously at ambient temperatures down to 25° C. or less, allow for massive multiplexing, high accuracy, minimal non-specific signal generation, and are easily reprogrammable.
The present disclosure relates to multiplex assay methods used to detect target nucleic acids of interest from several to many sources in a sample without amplification of the target nucleic acids of interest. The methods involve partitioning of first ribonucleoprotein complexes to detect short sequences from several to many loci scattered throughout a single source genome, and the addition of signal boosting second ribonucleoprotein complexes and reporter moieties.
The present disclosure relates to multiplex assay methods used to detect target nucleic acids of interest from several to many sources in a sample without amplification of the target nucleic acids of interest. The methods involve partitioning of first ribonucleoprotein complexes to detect short sequences from several to many loci scattered throughout a single source genome, and the addition of signal boosting second ribonucleoprotein complexes and reporter moieties.
The present disclosure relates to variant engineered nucleic acid-guided nucleases that may be used in CRISPR-based cascade assay systems to detect one or more target nucleic acids in a sample. The variant nucleases comprise an activity such that double-stranded DNA substrates do not bind to or are not cleaved by variant LbCas12a nuclease, or bind to or are cleaved very slowly by the variant nuclease, however single-stranded DNA substrates can bind and are cleaved by the variant nuclease, and wherein the variant nuclease exhibits both cis- and trans-cleavage activity.
The present disclosure relates to compositions of matter and assay methods used to detect one or more target nucleic acids of interest in a sample. The compositions and methods provide signal boost upon detection of target nucleic acids of interest in less than one minute and in some instances instantaneously at ambient temperatures down to 16° C. or less, without amplification of the target nucleic acids yet allowing for massive multiplexing, high accuracy and minimal non-specific signal generation.
The Board of Trustees of the University of Illinois (USA)
Inventor
Pandey, Ashish
Ganguli, Anurup
Mostafa, Ariana
Berger, Jacob
Abstract
The present disclosure describes compositions of matter comprising a ribonucleoprotein complex comprising a nucleic acid-guided nuclease and a guide RNA, and further comprising a blocking nucleic acid molecule represented by Formula IV, wherein Formula IV in the 5′-to-3′ direction comprises: T-D-M-A-Lp-C; wherein T is 17-31 nucleotides in length; D is 0-15 nucleotides in length; M is 1-25 nucleotides in length; A is 0-15 nucleotides in length and comprises a sequence complementary to D; and L is 3-25 nucleotides in length; p is 0 or 1; C is 4-15 nucleotides in length and comprises a sequence complementary to T; and wherein the blocking nucleic acid molecule comprises a sequence complementary to a gRNA.
The present disclosure relates to compositions of matter and assay methods used to detect one or more target nucleic acids of interest in a sample. The compositions and methods allow one to control reaction kinetics of the cascade assay by two orders of magnitude via molecular design of one of the reaction components; further, varying molecular design also allows for quantification of target nucleic acids of interest over a large range of concentrations or discriminating between extremely low copy numbers of target nucleic acids of interest.
The present disclosure relates to compositions of matter and assay methods used to detect one or more non-nucleic acid targets of interest in a sample. The compositions and methods provide signal boost upon detection of non-nucleic acid targets of interest in less than one minute and in some instances instantaneously at ambient temperatures down to 25° C. or less, allow for massive multiplexing, high accuracy, minimal non-specific signal generation, and are easily reprogrammable.
The present disclosure relates to compositions of matter and assay methods used to detect one or more non-nucleic acid targets of interest in a sample. The compositions and methods provide signal boost upon detection of non-nucleic acid targets of interest in less than one minute and in some instances instantaneously at ambient temperatures down to 25°C or less, allow for massive multiplexing, high accuracy, minimal non-specific signal generation, and are easily reprogrammable.
The present disclosure relates to compositions of matter and assay methods used to screen for molecular dimerization events using a two-ribonucleoprotein complex signal boost assay. The compositions and methods provide a readout upon detection of dimerization of molecules and may be implemented in a high throughput manner using libraries of tens to hundreds to thousands of putative binding partners.
The present disclosure relates to compositions of matter and methods used to activate effector nucleic acids and effector targets in vivo via a CRISPR-based cascade system. The compositions and methods achieve non-specific delivery of cascade system components to cells yet the cascade system works in a cell-specific manner.
A61K 31/7105 - Natural ribonucleic acids, i.e. containing only riboses attached to adenine, guanine, cytosine or uracil and having 3'-5' phosphodiester links
C12N 15/10 - Processes for the isolation, preparation or purification of DNA or RNA
C12N 15/90 - Stable introduction of foreign DNA into chromosome
The present disclosure relates to compositions of matter and assay methods used to screen for molecular dimerization events using a two-ribonucleoprotein complex signal boost assay. The compositions and methods provide a readout upon detection of dimerization of molecules and may be implemented in a high throughput manner using libraries of tens to hundreds to thousands of putative binding partners.
The present disclosure relates to compositions of matter and assay methods used to detect one or more target nucleic acids of interest in a sample. The compositions and methods allow one to control reaction kinetics of the cascade assay by two orders of magnitude via molecular design of one of the reaction components; further, varying molecular design also allows for quantification of target nucleic acids of interest over a large range of concentrations or discriminating between extremely low copy numbers of target nucleic acids of interest.
The present disclosure relates to compositions of matter and assay methods used to detect one or more target nucleic acids of interest in a sample. The compositions and methods provide signal boost upon detection of target nucleic acids of interest in less than one minute and in some instances instantaneously at ambient temperatures down to 16° C. or less, without amplification of the target nucleic acids yet allowing for massive multiplexing, high accuracy and minimal non-specific signal generation.
The Board of Trustees of The University of Illinois (USA)
Inventor
Ganguli, Anurup
Mostafa, Ariana
Berger, Jacob
Pandey, Ashish
Abstract
The present disclosure describes compositions of matter comprising a ribonucleoprotein complex comprising a nucleic acid-guided nuclease and a guide RNA, and further comprising a blocking nucleic acid molecule represented by Formula IV, wherein Formula IV in the 5′-to-3′ direction comprises: T-D-M-A-Lp-C; wherein T is 17-31 nucleotides in length; D is 0-15 nucleotides in length; M is 1-25 nucleotides in length; A is 0-15 nucleotides in length and comprises a sequence complementary to D; and L is 3-25 nucleotides in length; p is 0 or 1; C is 4-15 nucleotides in length and comprises a sequence complementary to T; and wherein the blocking nucleic acid molecule comprises a sequence complementary to a gRNA.
The present disclosure relates to compositions of matter and assay methods used to detect one or more target nucleic acids of interest in a sample. The compositions and methods allow one to control reaction kinetics of the cascade assay by two orders of magnitude via molecular design of one of the reaction components; further, varying molecular design also allows for quantification of target nucleic acids of interest over a large range of concentrations or discriminating between extremely low copy numbers of target nucleic acids of interest.
J-C are separate nucleic acid strands; A is 0-15 nucleotides in length and comprises at least 50% sequence complementarity to D; B is 4-12 nucleotides in length and comprises at least 50% sequence complementarity to T; L is 3-25 nucleotides in length; J is an integer between 1 and 10; C is 4-15 nucleotides in length; and wherein the blocking nucleic acid molecule comprises a sequence complementary to a gRNA.
The Board of Trustees of The University of Illinois (USA)
Inventor
Ganguli, Anurup
Mostafa, Ariana
Berger, Jacob
Pandey, Ashish
Bashir, Rashid
Abstract
J-C and T-D are separate nucleic acid strands; T is 17-135 nucleotides in length and comprises at least 50% sequence complementarity to B and C; D is 0-10 nucleotides in length and comprises at least 50% sequence complementarity to A; and wherein the blocking nucleic acid molecule comprises a sequence complementary to a gRNA.
The present disclosure relates to compositions of matter and assay methods used to detect one or more target nucleic acids of interest in a sample. The compositions and methods allow one to control reaction kinetics of the cascade assay by two orders of magnitude via molecular design of one of the reaction components; further, varying molecular design also allows for quantification of target nucleic acids of interest over a large range of concentrations or discriminating between extremely low copy numbers of target nucleic acids of interest.
The present disclosure relates to compositions of matter and assay methods used to detect one or more target nucleic acids of interest in a sample. The compositions and methods provide signal boost upon detection of target nucleic acids of interest in less than one minute and in some instances instantaneously at ambient temperatures down to 16° C. or less, without amplification of the target nucleic acids yet allowing for massive multiplexing, high accuracy and minimal non-specific signal generation.
01 - Chemical and biological materials for industrial, scientific and agricultural use
05 - Pharmaceutical, veterinary and sanitary products
10 - Medical apparatus and instruments
Goods & Services
Diagnostic preparations for scientific or research use; Diagnostic preparations for clinical research purposes; Diagnostic reagents for testing proteins, DNA and RNA samples for scientific use Kits comprised of medical diagnostic reagents and assays for testing proteins, DNA and RNA samples for medical purposes Medical diagnostic apparatus for testing proteins, DNA and RNA samples
01 - Chemical and biological materials for industrial, scientific and agricultural use
05 - Pharmaceutical, veterinary and sanitary products
10 - Medical apparatus and instruments
Goods & Services
Diagnostic preparations for scientific or research use; Diagnostic preparations for clinical research purposes; Diagnostic reagents for testing proteins, DNA and RNA samples for scientific use Kits comprised of medical diagnostic reagents and assays for testing proteins, DNA and RNA samples for medical purposes Medical diagnostic apparatus for testing proteins, DNA and RNA samples
40.
Tuning cascade assay kinetics via molecular design
The present disclosure relates to compositions of matter and assay methods used to detect one or more target nucleic acids of interest in a sample. The compositions and methods allow one to control reaction kinetics of the cascade assay by two orders of magnitude via molecular design of one of the reaction components; further, varying molecular design also allows for quantification of target nucleic acids of interest over a large range of concentrations or discriminating between extremely low copy numbers of target nucleic acids of interest.
The present disclosure relates to compositions of matter and assay methods used to detect one or more target nucleic acids of interest in a sample. The compositions and methods provide signal boost upon detection of target nucleic acids of interest in less than one minute and in some instances instantaneously at ambient temperatures down to 16° C. or less, without amplification of the target nucleic acids yet allowing for massive multiplexing, high accuracy and minimal non-specific signal generation.
01 - Chemical and biological materials for industrial, scientific and agricultural use
05 - Pharmaceutical, veterinary and sanitary products
10 - Medical apparatus and instruments
Goods & Services
Diagnostic preparations for scientific or research use; Diagnostic preparations for clinical research purposes; Diagnostic reagents for testing proteins, DNA and RNA samples for scientific use Kits comprised of medical diagnostic reagents and assays for testing proteins, DNA and RNA samples for medical purposes Medical diagnostic apparatus for testing proteins, DNA and RNA samples
01 - Chemical and biological materials for industrial, scientific and agricultural use
05 - Pharmaceutical, veterinary and sanitary products
10 - Medical apparatus and instruments
Goods & Services
Diagnostic reagents and kits for testing proteins, DNA and RNA samples for scientific use Diagnostic reagents and kits for testing proteins, DNA and RNA samples for medical purposes; Diagnostic preparations for medical laboratory use; Medical diagnostic apparatus for testing DNA and RNA samples
J-C and T-D are separate nucleic acid strands; T is 17-135 nucleotides in length and comprises at least 50% sequence complementarity to B and C; D is 0-10 nucleotides in length and comprises at least 50% sequence complementarity to A; and wherein the blocking nucleic acid molecule comprises a sequence complementary to a gRNA.
The present disclosure describes a nucleic acid-guided nuclease cascade assay that can detect one or more target nucleic acids of interest of interest at attamolar (aM) (or lower) limits in about 10 minutes or less without the need for amplifying the target nucleic acids of interest. The nucleic acid-guided nuclease cascade assays utilize signal amplification mechanisms comprising various components including nucleic acid-guided nucleases, guide RNAs (gRNAs), blocked nucleic acid molecules, blocked primer molecules, and reporter moieties.
