Photoblocked probes are disclosed including a specific hydrolysis probe having a first nucleic acid sequence complementary to a target having a second nucleic acid sequence, a first and a second interactive labels, a 5′ end and a 3′ end, and one or more photocleavable moieties coupled to one or more nucleotides of the specific hydrolysis probe, wherein the photocleaveable moiety interferes with the hybridization of the specific hydrolysis probe with the region of the amplification product. Also disclosed are PCR methods for the detection of the presence or absence of a target nucleic acid in a sample utilizing the photoblocked probes, as well as kits.

Photoblocked probes are disclosed including a specific hydrolysis probe having a first nucleic acid sequence complementary to a target having a second nucleic acid sequence, a first and a second interactive labels, a 5′ end and a 3′ end, and one or more photocleavable moieties coupled to one or more nucleotides of the specific hydrolysis probe, wherein the photocleaveable moiety interferes with the hybridization of the specific hydrolysis probe with the region of the amplification product. Also disclosed are PCR methods for the detection of the presence or absence of a target nucleic acid in a sample utilizing the photoblocked probes, as well as kits.

Methods of generating a nucleic acid signature for identifying particles associated in a partition are provided. In one aspect, the method comprises: partitioning a sample into a plurality of partitions comprising a particle comprising a solid support surface, the solid support surface having a plurality of oligonucleotide primers conjugated thereon, wherein the oligonucleotide primers comprise a barcode sequence, and wherein the partitions have 0, 1, or more than 1 particles per partition; providing in a partition a substrate comprising a barcode sequence or repeating clonal barcode sequences; and in the partition, associating a first particle conjugated to oligonucleotide primers comprising a first barcode sequence and a second particle conjugated to oligonucleotide primers comprising a second barcode sequence to a barcode sequence from the substrate, thereby generating a nucleic acid signature for the particles in the partition.

Methods of generating a nucleic acid signature for identifying particles associated in a partition are provided. In one aspect, the method comprises: partitioning a sample into a plurality of partitions comprising a particle comprising a solid support surface, the solid support surface having a plurality of oligonucleotide primers conjugated thereon, wherein the oligonucleotide primers comprise a barcode sequence, and wherein the partitions have 0, 1, or more than 1 particles per partition; providing in a partition a substrate comprising a barcode sequence or repeating clonal barcode sequences; and in the partition, associating a first particle conjugated to oligonucleotide primers comprising a first barcode sequence and a second particle conjugated to oligonucleotide primers comprising a second barcode sequence to a barcode sequence from the substrate, thereby generating a nucleic acid signature for the particles in the partition.

Methods and systems for the high throughput RNA profiling or sequencing analysis of fixed single cells are provided. The methods and systems utilize barcoded microcarriers comprising a small carrier bead attached to a plurality of molecular barcodes with cleavable linkers, wherein these barcoded microcarriers are then combined in a system (e.g., a sealed system) with a single cell. The molecular barcodes are then released from the carrier beads to anneal to the mRNAs in the cells. Using reverse transcription, barcoded cDNAs is then generated from the mRNAs and the cells are lysed or digested to release the barcoded cDNAs for sequencing.

Methods and systems for the high throughput RNA profiling or sequencing analysis of fixed single cells are provided. The methods and systems utilize barcoded microcarriers comprising a small carrier bead attached to a plurality of molecular barcodes with cleavable linkers, wherein these barcoded microcarriers are then combined in a system (e.g., a sealed system) with a single cell. The molecular barcodes are then released from the carrier beads to anneal to the mRNAs in the cells. Using reverse transcription, barcoded cDNAs is then generated from the mRNAs and the cells are lysed or digested to release the barcoded cDNAs for sequencing.

The invention is directed to a method of identifying the interactions of RNA such as miRNA with small molecules interacting with RNA (SMIRNAs). A candidate SMIRNA group is associated with a photoaffinity diazirene group that can form a covalent complex with an RNA target site and with an alkyne group that can be used in subsequent “click chemistry’ reactions such as a “CuAAC” reaction, a copper-catalyzed alkyne-azide cy-cloaddition to yield a stable triazole ring. By this means, the RNA binding site of the small molecule can be identified via isolation of the RNA-targetSMIRNA covalent complex and reverse transcription of the RNA followed by DNA sequencing of the reverse transcription product. Sites on the RNA blocked during reverse transcription by the covalently bound SMIRNA are identified as terminations in the sequence compared to the native RNA.

The invention is directed to a method of identifying the interactions of RNA such as miRNA with small molecules interacting with RNA (SMIRNAs). A candidate SMIRNA group is associated with a photoaffinity diazirene group that can form a covalent complex with an RNA target site and with an alkyne group that can be used in subsequent “click chemistry’ reactions such as a “CuAAC” reaction, a copper-catalyzed alkyne-azide cy-cloaddition to yield a stable triazole ring. By this means, the RNA binding site of the small molecule can be identified via isolation of the RNA-targetSMIRNA covalent complex and reverse transcription of the RNA followed by DNA sequencing of the reverse transcription product. Sites on the RNA blocked during reverse transcription by the covalently bound SMIRNA are identified as terminations in the sequence compared to the native RNA.

Provided are methods and systems useful for screening large libraries of effector molecules. Such methods and systems are particularly useful in microfluidic systems and devices. The methods and systems provided herein utilize encoded effectors to screen large libraries of effectors.

Provided are methods and systems useful for screening large libraries of effector molecules. Such methods and systems are particularly useful in microfluidic systems and devices. The methods and systems provided herein utilize encoded effectors to screen large libraries of effectors.