Abstract: The present disclosure provides a method for enrichment of at least one target nucleic acid in a library of nucleic acids. A first oligonucleotide is hybridized to a target nucleic acid in library of nucleic acids having first and second adapters. The hybridized first oligonucleotide is extended with a first polymerase, thereby producing a first primer extension complex including the target nucleic acid and the extended first oligonucleotide. The first primer extension complex is captured, enriched relative to the library of nucleic acids, and a second oligonucleotide is hybridized to the target nucleic acid. The hybridized second oligonucleotide is extended with a second polymerase, thereby producing a second primer extension complex including the target nucleic acid and the extended second oligonucleotide, and further liberating the extended first oligonucleotide from the first primer extension complex.

Abstract: The invention is a novel method of sequencing nucleic acids involving making and sequencing a library of single stranded circular target nucleic acids.

Abstract: Disclosed herein are methods and compositions for normalizing polynucleotide concentration. Normalizing may be accomplished using polynucleotide binding proteins (e.g., catalytically inactive CRISPR protein or catalytically inactive Argonaute proteins). The polynucleotide binding proteins may bind to the adapter sequences of a target polynucleotide. Thus, adding the same amount of a polynucleotide binding proteins to different samples and then extracting the polynucleotide protein is shown herein to extract similar amounts of target polynucleotides from the different samples.

Abstract: The invention is a novel method of sequencing nucleic acids involving making and sequencing a library of double stranded target nucleic acids containing a linear partially single-stranded adaptor.

Abstract: Disclosed herein are methods and compositions for normalizing polynucleotide concentration. Normalizing may be accomplished using polynucleotide binding proteins (e.g., catalytically inactive CRISPR protein or catalytically inactive Argonaute proteins). The polynucleotide binding proteins may bind to the adapter sequences of a target polynucleotide. Thus, adding the same amount of a polynucleotide binding proteins to different samples and then extracting the polynucleotide protein is shown herein to extract similar amounts of target polynucleotides from the different samples.

Abstract: Disclosed is a method of fragmenting DNA comprising contacting a sample of target DNA with (a) a composition comprising an active transpososome, and (b) a composition comprising an inactive transpososome, under conditions suitable for transpososome activity, wherein a ratio of an amount of the inactive transpososome in the composition of (b) to an amount of the active transpososome in the composition of (a) determines the mean fragment size and a level of insertion bias.

Abstract: The invention is a novel method of separately sequencing each strand of a nucleic acid involving the use of an adaptor comprising a strand cleavage site or a strand synthesis termination site. The adaptor may also be self-priming at the strand cleavage site.

Abstract: The invention is a method of single cell transcriptome analysis. The method comprises detecting multiple transcripts in each individual cell of the plurality of cells by barcoding the transcripts with a cell-specific compound barcode formed using a DNA polymerase and a terminal transferase, optionally in a single enzyme such as a reverse transcriptase.

Abstract: The invention provides sets of RNA depletion probes, short DNA oligos that hybridize along the length of a target RNA and mediate digestion of the target RNA by RNase H to remove super-abundant RNA molecules from a sample. Depletion probes according to the invention are designed foremost based on biochemistry and the biophysical properties of the probes so that all of the depletion probes of a set exhibit substantially uniform, consistent behavior in binding to a target RNA in a sample. Probes are principally designed to specific performance targets and biophysical properties, yielding probe sets with irregular, even apparently random, spacing along a target RNA molecule.

Abstract: The invention provides sets of RNA depletion probes, short DNA oligos that hybridize along the length of a target RNA and mediate digestion of the target RNA by RNase H to remove super-abundant RNA molecules from a sample. Depletion probes according to the invention are designed foremost based on biochemistry and the biophysical properties of the probes so that all of the depletion probes of a set exhibit substantially uniform, consistent behavior in binding to a target RNA in a sample. Probes are principally designed to specific performance targets and biophysical properties, yielding probe sets with irregular, even apparently random, spacing along a target RNA molecule.

Abstract: The invention is a novel method of separately sequencing each strand of a nucleic acid involving the use of an adaptor comprising a strand cleavage site or a strand synthesis termination site. The adaptor may also be self-priming at the strand cleavage site.

Abstract: The disclosure provides a composition comprising a double-stranded deoxyribonucleic acid (dsDNA) sequence comprising from 5? to 3?, a sequence comprising a first adaptor sequence, a template sequence, and a second adaptor sequence, wherein the second adaptor sequence comprises a hybridization site for a template switching oligonucleotide (TSO). The disclosure provides methods for making the compositions of the disclosure using a template switching mechanism to add non-templated basepairs to the ends of a DNA molecule, hybridize a TSO to the non-templated basepairs, and then extend the sequence complementary to the TSO to add an adaptor.

Abstract: The disclosure provides a composition comprising a double-stranded deoxyribonucleic acid (dsDNA) sequence comprising from 5? to 3?, a sequence comprising a first adaptor sequence, a template sequence, and a second adaptor sequence, wherein the second adaptor sequence comprises a hybridization site for a template switching oligonucleotide (TSO). The disclosure provides methods for making the compositions of the disclosure using a template switching mechanism to add non-templated basepairs to the ends of a DNA molecule, hybridize a TSO to the non-templated basepairs, and then extend the sequence complementary to the TSO to add an adaptor.

Abstract: The invention is a method of single cell transcriptome analysis. The method comprises detecting multiple transcripts in each individual cell of the plurality of cells by barcoding the transcripts with a cell-specific compound barcode formed using a DNA polymerase and a terminal transferase, optionally in a single enzyme such as a reverse transcriptase.

Abstract: The invention is a novel method of making and using a library such as a sequencing library of single stranded circular nucleic acid templates via splint ligation. In particular, disclosed are methods of making circular target nucleic acid molecules and libraries of such molecules for downstream analysis such as nucleic acid sequencing. The method comprises the steps of adding universal sequences to nucleic acid molecules, rendering single-stranded these nucleic acid molecules with universal sequences on their ends by contacting with a probe complementary to at least a portion of the universal sequences, and allowing the hybridized probe to enable circularization and formation of single-stranded circular (sscDNA) molecules.

Abstract: The invention is a novel method of sequencing nucleic acids involving making and sequencing a library of double stranded target nucleic acids containing a linear partially single-stranded adaptor.

Abstract: The present invention provides synthetic nucleic acid molecule tags that can be added into samples for identification and tracking. Among other things, the present invention provides synthetic, high-molecular weight concatemers, which can be combined with samples to yield hundreds of millions of unique identifiers. Examples applications for which the synthetic nucleic acid molecule tags can be used, include industrial, research and clinical applications.

Abstract: The invention is a novel method of sequencing nucleic acids involving making and sequencing a library of single stranded circular target nucleic acids.

Abstract: The disclosure provides compositions and methods for high-throughput screening a plurality of transposases to identify rare mutations that affect desired features of the transposase. Compositions of the disclosures are provided that facilitate the high-throughput screening methods of the disclosure.

Abstract: The invention is a novel method of making and using a library such as a sequencing library of single stranded circular nucleic acid templates via splint ligation.