An example of a kit includes a flow cell, a primer fluid, and a cleaving fluid. The flow cell includes at least one surface functionalized with a polymeric hydrogel including azide functional groups or amine functional groups. The primer fluid includes a plurality of alkyne-containing primers, each alkyne-containing primer having an amino cleavable group attaching a primer sequence of the alkyne-containing primer to an alkyne-containing moiety of the alkyne-containing primer. The cleaving fluid includes a substance that is reactive with the amino cleavable group.

An example of a kit includes a flow cell, a primer fluid, and a cleaving fluid. The flow cell includes at least one surface functionalized with a polymeric hydrogel including azide functional groups or amine functional groups. The primer fluid includes a plurality of alkyne-containing primers, each alkyne-containing primer having an amino cleavable group attaching a primer sequence of the alkyne-containing primer to an alkyne-containing moiety of the alkyne-containing primer. The cleaving fluid includes a substance that is reactive with the amino cleavable group.

The present invention is directed to methods and compositions for generating a pool of oligonucleotides. The invention finds use in preparing a population or subpopulations of oligonucleotides in solution. The pool of oligonucleotides finds use in a variety of nucleic acid detection and/or amplification assays.

The present invention is directed to methods and compositions for generating a pool of oligonucleotides. The invention finds use in preparing a population or subpopulations of oligonucleotides in solution. The pool of oligonucleotides finds use in a variety of nucleic acid detection and/or amplification assays.

The present disclosure relates to compositions, methods, and kits for generating capture probes on a substrate for identifying the location of analytes in a biological sample. In particular, disclosed is a method of generating a spatial array comprising: (a) providing a substrate comprising a plurality of acceptor oligonucleotides, wherein an acceptor oligonucleotide of the plurality of acceptor oligonucleotides comprises a spatial barcode and a first ligation handle, and wherein the 5′ end of the acceptor oligonucleotide is attached to the substrate; (b) providing a plurality of universal splint oligonucleotides, wherein a universal splint oligonucleotide of the plurality of universal splint oligonucleotides comprises a sequence complementary to the first ligation handle and a sequence complementary to a second ligation handle present in a donor oligonucleotide of a plurality of donor oligonucleotides; and (c) ligating the donor oligonucleotide comprising a capture domain to the 3′ end of the acceptor oligonucleotide to generate a capture probe, wherein the universal splint oligonucleotide is hybridized to the first ligation handle and the second ligation handle, thereby generating a spatial array.

The present disclosure relates to compositions, methods, and kits for generating capture probes on a substrate for identifying the location of analytes in a biological sample. In particular, disclosed is a method of generating a spatial array comprising: (a) providing a substrate comprising a plurality of acceptor oligonucleotides, wherein an acceptor oligonucleotide of the plurality of acceptor oligonucleotides comprises a spatial barcode and a first ligation handle, and wherein the 5′ end of the acceptor oligonucleotide is attached to the substrate; (b) providing a plurality of universal splint oligonucleotides, wherein a universal splint oligonucleotide of the plurality of universal splint oligonucleotides comprises a sequence complementary to the first ligation handle and a sequence complementary to a second ligation handle present in a donor oligonucleotide of a plurality of donor oligonucleotides; and (c) ligating the donor oligonucleotide comprising a capture domain to the 3′ end of the acceptor oligonucleotide to generate a capture probe, wherein the universal splint oligonucleotide is hybridized to the first ligation handle and the second ligation handle, thereby generating a spatial array.

The invention provides a method for detecting the presence of a target nucleic acid analyte, for example a pathogen or virus nucleic acid, in a sample using oligonucleotide probe-functionalised nanoparticles, where hybridisation of at least three different oligonucleotide probes to at least three different target sequences in the target analyte causes agglomeration of the nanoparticles and a visible colour change. The invention also provides a population of such oligonucleotide probe-functionalised nanoparticles and a related kit for detection of a target nucleic acid analyte.

The invention provides a method for detecting the presence of a target nucleic acid analyte, for example a pathogen or virus nucleic acid, in a sample using oligonucleotide probe-functionalised nanoparticles, where hybridisation of at least three different oligonucleotide probes to at least three different target sequences in the target analyte causes agglomeration of the nanoparticles and a visible colour change. The invention also provides a population of such oligonucleotide probe-functionalised nanoparticles and a related kit for detection of a target nucleic acid analyte.

The present application provides methods for detecting a target nucleic acid molecule in a sample comprising contacting said sample with a ligatable probe comprising one or more parts and allowing said probe to hybridise to the target nucleic acid molecule, ligating any probe which has hybridised to the target nucleic acid molecule, amplifying the ligated probe, and detecting the amplification product, thereby to detect the target nucleic acid molecule, wherein said probes comprise at least one ribonucleotide at or near to a ligation site and/or wherein the probe or a probe part comprises an additional sequence 5′ to a target-specific binding site which is not hybridised to the target nucleic acid molecule upon hybridisation of the probe to the target nucleic acid molecule and forms a 5′ flap containing one or more nucleotides at its 3′ end that is cleaved prior to ligation, and methods of synthesising a DNA molecule with Phi29 DNA polymerase using a template nucleic acid molecule comprising at least one ribonucleotide. Probes for use in the detection methods are provided.

The present application provides methods for detecting a target nucleic acid molecule in a sample comprising contacting said sample with a ligatable probe comprising one or more parts and allowing said probe to hybridise to the target nucleic acid molecule, ligating any probe which has hybridised to the target nucleic acid molecule, amplifying the ligated probe, and detecting the amplification product, thereby to detect the target nucleic acid molecule, wherein said probes comprise at least one ribonucleotide at or near to a ligation site and/or wherein the probe or a probe part comprises an additional sequence 5′ to a target-specific binding site which is not hybridised to the target nucleic acid molecule upon hybridisation of the probe to the target nucleic acid molecule and forms a 5′ flap containing one or more nucleotides at its 3′ end that is cleaved prior to ligation, and methods of synthesising a DNA molecule with Phi29 DNA polymerase using a template nucleic acid molecule comprising at least one ribonucleotide. Probes for use in the detection methods are provided.