This disclosure provides, inter alia, a probe system probe system for analyzing a nucleic acid sample. In some embodiments, the probe system may comprise: a set of identifier oligonucleotides of sequence B, a set of splint oligonucleotides of formula X-A-B-Z, wherein sequence A is complementary to a genomic fragment and sequence B is complementary to at least one member of the set of identifier oligonucleotides, and one or more probe sequences comprising X and Z. Each splint oligonucleotide is capable of hybridizing to the probe sequences, a member of the set of identifier oligonucleotides and a genomic fragment, thereby producing a ligatable complex of formula X-A-B-Z. The probe system can be used to identify a chromosome aneuploidy in cell free DNA, for example.

This disclosure provides, inter alia, a probe system probe system for analyzing a nucleic acid sample. In some embodiments, the probe system may comprise: a set of identifier oligonucleotides of sequence B, a set of splint oligonucleotides of formula X-A-B-Z, wherein sequence A is complementary to a genomic fragment and sequence B is complementary to at least one member of the set of identifier oligonucleotides, and one or more probe sequences comprising X and Z. Each splint oligonucleotide is capable of hybridizing to the probe sequences, a member of the set of identifier oligonucleotides and a genomic fragment, thereby producing a ligatable complex of formula X-A-B-Z. The probe system can be used to identify a chromosome aneuploidy in cell free DNA, for example.

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.

Compositions and reagents for molecular profiling using proximity ligation-/>7 situ hybridization (PLISH) are disclosed. In particular, PLISH merges the specificity of proximity ligation, the sensitivity of tiling multiple probes for a target nucleic acid, and the high signal intensity of rolling circle amplification. The probe design capitalizes on the formation of Holliday-like junctions for optimal signal amplification. PLISH provides single molecule resolution and allows for quantitation of a virtually unlimited number of nucleic acids within individual cells. PLISH is also compatible with immunohistochemistry and archival formal-fixed, paraffin-embedded tissue samples.

Compositions and reagents for molecular profiling using proximity ligation-/>7 situ hybridization (PLISH) are disclosed. In particular, PLISH merges the specificity of proximity ligation, the sensitivity of tiling multiple probes for a target nucleic acid, and the high signal intensity of rolling circle amplification. The probe design capitalizes on the formation of Holliday-like junctions for optimal signal amplification. PLISH provides single molecule resolution and allows for quantitation of a virtually unlimited number of nucleic acids within individual cells. PLISH is also compatible with immunohistochemistry and archival formal-fixed, paraffin-embedded tissue samples.

Provided herein is a multi-sample and multi-locus method for analyzing a genetic locus. In particular, provided herein is a method for SNP detection and analysis based on high-throughput sequencing, comprising designing a probe, pre-amplification and biotin labeling, hybridization, ligation, barcode specific primer extension, sequencing and analyzing the SNP locus. A probe set is for the analysis is also provided.

Provided herein is a multi-sample and multi-locus method for analyzing a genetic locus. In particular, provided herein is a method for SNP detection and analysis based on high-throughput sequencing, comprising designing a probe, pre-amplification and biotin labeling, hybridization, ligation, barcode specific primer extension, sequencing and analyzing the SNP locus. A probe set is for the analysis is also provided.

Ligation assays in liquid phase for detecting nucleic acid sequences.

Ligation assays in liquid phase for detecting nucleic acid sequences.