The present disclosure relates to a method of sequencing nascent RNA in a cell. In some embodiments, the nascent RNA is conjugated to DNA using copper-catalyzed azide-alkyne cycloaddition (CuAAC). Methods of the present disclosure can be used to generate genomic libraries of a cell and measure gene expression and enhancer and/or super-enhancer activity.

Disclosed here is a method for detecting genome rearrangement in a biological sample, comprising: obtaining a contact matrix plotted from proximity ligation sequencing data of at least one chromosome; identifying an abnormal contact pattern in the contact matrix compared to the contact matrix of a reference genome; comparing the abnormal contact pattern in the contact matrix to one or more known patterns associated with genomic rearrangement to identify a type of genomic rearrangement causing the abnormal contact pattern. Also disclosed is a method for detecting genome rearrangement in a biological sample, comprising: selecting linked chromosomal fragments from proximity ligation sequencing data of at least one chromosome, identifying an abnormal covalent bonding pattern of the linked chromosomal fragments compared to a reference genome; and comparing the abnormal covalent bonding pattern to one or more known patterns associated with genomic rearrangement to identify genomic rearrangement causing the abnormal covalent bonding pattern.

Disclosed here is a method for detecting genome rearrangement in a biological sample, comprising: obtaining a contact matrix plotted from proximity ligation sequencing data of at least one chromosome; identifying an abnormal contact pattern in the contact matrix compared to the contact matrix of a reference genome; comparing the abnormal contact pattern in the contact matrix to one or more known patterns associated with genomic rearrangement to identify a type of genomic rearrangement causing the abnormal contact pattern. Also disclosed is a method for detecting genome rearrangement in a biological sample, comprising: selecting linked chromosomal fragments from proximity ligation sequencing data of at least one chromosome, identifying an abnormal covalent bonding pattern of the linked chromosomal fragments compared to a reference genome; and comparing the abnormal covalent bonding pattern to one or more known patterns associated with genomic rearrangement to identify genomic rearrangement causing the abnormal covalent bonding pattern.

An inventive method for amplifying at least one RNA which is contained in a sample includes reverse transcribing the at least one RNA using a first primer, adding a dideoxy nucleotide which is modified in the 3′-position with a first partner of a pair of azide and alkyne molecules by action of a template independent polymerase to attach a single 3′-azide- or 3′-alkyne-modified dideoxy nucleotide at the 3′-end of the obtained cDNA, adding an adapter molecule which comprises a polynucleotide sequence and at its 5′-end a second partner of such pair of azide and alkyne molecules and ligating the adapter to the 3′-modified cDNA under formation of a triazole linkage, adding a second primer which is complementary to at least a part of the adapter molecule and which contains at its 3′-end a nucleotide which is complementary to the dideoxy nucleotide at the 3′-end of the cDNA to effect hybridization and binding of the second primer overlapping the triazole linkage, adding a third primer and amplifying the full length cDNA. Variations of this method are also disclosed. Uses of such method especially for preparing a full length RNA library and for sequencing of a plurality of RNAs contained in a sample, as well as reagent kits for performing such methods are also disclosed and included in the invention.

An inventive method for amplifying at least one RNA which is contained in a sample includes reverse transcribing the at least one RNA using a first primer, adding a dideoxy nucleotide which is modified in the 3′-position with a first partner of a pair of azide and alkyne molecules by action of a template independent polymerase to attach a single 3′-azide- or 3′-alkyne-modified dideoxy nucleotide at the 3′-end of the obtained cDNA, adding an adapter molecule which comprises a polynucleotide sequence and at its 5′-end a second partner of such pair of azide and alkyne molecules and ligating the adapter to the 3′-modified cDNA under formation of a triazole linkage, adding a second primer which is complementary to at least a part of the adapter molecule and which contains at its 3′-end a nucleotide which is complementary to the dideoxy nucleotide at the 3′-end of the cDNA to effect hybridization and binding of the second primer overlapping the triazole linkage, adding a third primer and amplifying the full length cDNA. Variations of this method are also disclosed. Uses of such method especially for preparing a full length RNA library and for sequencing of a plurality of RNAs contained in a sample, as well as reagent kits for performing such methods are also disclosed and included in the invention.

Sensitive and specific detection of nucleic acids can be achieved using a chemical ligation-based template assisted rapid assay (TARA-L) with simple chemical reactions between probes and without the need for enzymes. Probes are designed to form a ligation product when they anneal to adjacent portions of a target nucleic acid. The ligation products can be detected, such as in immunochromatographic assays. The methods allow for the fast, efficient analysis of biological samples for the presence of nucleic acids and can be used, for example, in point of care settings.

Sensitive and specific detection of nucleic acids can be achieved using a chemical ligation-based template assisted rapid assay (TARA-L) with simple chemical reactions between probes and without the need for enzymes. Probes are designed to form a ligation product when they anneal to adjacent portions of a target nucleic acid. The ligation products can be detected, such as in immunochromatographic assays. The methods allow for the fast, efficient analysis of biological samples for the presence of nucleic acids and can be used, for example, in point of care settings.

In certain aspects, the invention disclosed herein relates to the isothermal amplification of probe linkage products to generate specific amplified signals. In some aspects, the invention provides methods, reagents, and kits for carrying out such amplification via the isothermal chain reaction (ICR).

In certain aspects, the invention disclosed herein relates to the isothermal amplification of probe linkage products to generate specific amplified signals. In some aspects, the invention provides methods, reagents, and kits for carrying out such amplification via the isothermal chain reaction (ICR).

The present invention provides compositions, apparatuses and methods for detecting one or more nucleic acid targets present in a sample. Methods of the invention include utilizing two or more ligation probes that reversibly bind a target nucleic acid in close proximity to each other and possess complementary reactive ligation moieties. When such probes have bound to the target in the proper orientation, they are able to undergo a spontaneous chemical ligation reaction that yields a ligation product that is directly detected or that is amplified to produce amplicons that are then detected. The present invention also provides methods to stabilize sample RNA so that degradation does not significantly affect the results of the analysis.