The present disclosure describes technologies that permit sensitive detection of nucleic acids of interest (i.e., nucleic acids whose nucleotide sequence is or includes a target sequence).

Disclosed herein are methods of long range target specific amplification and sequencing using an RNA intermediate synthesized directly from the target including using hairpin adaptors having a double stranded promoter and an overhang which hybridizes with a reverse-complementary overhang on a target nucleic acid. RNA transcription eliminates clonal amplification of early synthesis errors. Approaches allow for the identification of target-adjacent sequence, such as sequence adjacent to a repeat element target. Also disclosed herein are compositions and kits for amplification and sequencing.

Disclosed herein are methods of long range target specific amplification and sequencing using an RNA intermediate synthesized directly from the target including using hairpin adaptors having a double stranded promoter and an overhang which hybridizes with a reverse-complementary overhang on a target nucleic acid. RNA transcription eliminates clonal amplification of early synthesis errors. Approaches allow for the identification of target-adjacent sequence, such as sequence adjacent to a repeat element target. Also disclosed herein are compositions and kits for amplification and sequencing.

Disclosed herein are methods of long range target specific amplification and sequencing using an RNA intermediate synthesized directly from the target including using hairpin adaptors having a double stranded promoter and an overhang which hybridizes with a reverse-complementary overhang on a target nucleic acid. RNA transcription eliminates clonal amplification of early synthesis errors. Approaches allow for the identification of target-adjacent sequence, such as sequence adjacent to a repeat element target. Also disclosed herein are compositions and kits for amplification and sequencing.

Provided herein are compositions and methods to assess the genomic landscape of fixed cells using light activated oligonucleotides that can be directed to the nucleus, mitochondria, or cytoplasm of fixed cells and that, upon activation, can be extended for in situ copying of nuclear single-stranded DNA (i.e., open chromatin), open mitochondrial DNA, and/or cytoplasmic RNA into barcoded complementary DNA. These methods also provide for gene specific 3D chromatin structural niche analysis.

Described herein are synthetic methods for producing sequence-specific RNA oligonucleotides that eliminate impurities produced in prior art methods. In one aspect, an end-protected capture DNA complementary to a portion of the product RNA is employed. In another aspect, the template DNA is covalently or noncovalently linked to the RNA polymerase, either directly or through the use of a nontemplate DNA. In a third aspect, a flow chamber is employed. All of the methods can be used in combination.

Described herein are synthetic methods for producing sequence-specific RNA oligonucleotides that eliminate impurities produced in prior art methods. In one aspect, an end-protected capture DNA complementary to a portion of the product RNA is employed. In another aspect, the template DNA is covalently or noncovalently linked to the RNA polymerase, either directly or through the use of a nontemplate DNA. In a third aspect, a flow chamber is employed. All of the methods can be used in combination.

Described herein are synthetic methods for producing sequence-specific RNA oligonucleotides that eliminate impurities produced in prior art methods. In one aspect, an end-protected capture DNA complementary to a portion of the product RNA is employed. In another aspect, the template DNA is covalently or noncovalently linked to the RNA polymerase, either directly or through the use of a nontemplate DNA. In a third aspect, a flow chamber is employed. All of the methods can be used in combination.

A method of RNA synthesis via fluid flow apparatus. The method may involve introducing into a first fluid flow module, via a plurality of inlet ports, a plurality of reactants comprising: at least one nucleoside triphosphate (NTP), a reaction buffer, and DNA, a DNA based compound or a DNA based mixture; allowing at least some of the reactants to react within a reaction channel or well within the first module of the flow system; retaining or recirculating the DNA at the first reactor module and allowing reaction products of the reactants to flow into a first fluidic filtration module; and filtering the reaction products within the first filtration module.

A method of RNA synthesis via fluid flow apparatus. The method may involve introducing into a first fluid flow module, via a plurality of inlet ports, a plurality of reactants comprising: at least one nucleoside triphosphate (NTP), a reaction buffer, and DNA, a DNA based compound or a DNA based mixture; allowing at least some of the reactants to react within a reaction channel or well within the first module of the flow system; retaining or recirculating the DNA at the first reactor module and allowing reaction products of the reactants to flow into a first fluidic filtration module; and filtering the reaction products within the first filtration module.