The invention relates to methods of enriching for target nucleic acid molecules, More particularly, the methods of enriching for target nucleic acid molecules comprise binding target nucleic acid molecules in a sample with one or more first target endonucleases that are specific to a first locus of a target region of the target nucleic acid molecules, separating the target nucleic acid molecules from nontarget nucleic acid molecules in the sample, and binding the separated target nucleic acid molecules with one or more second target endonucleases that are specific to a second locus of the target region of the target nucleic acid molecules, and uses thereof.

The invention relates to methods of enriching for target nucleic acid molecules, More particularly, the methods of enriching for target nucleic acid molecules comprise binding target nucleic acid molecules in a sample with one or more first target endonucleases that are specific to a first locus of a target region of the target nucleic acid molecules, separating the target nucleic acid molecules from nontarget nucleic acid molecules in the sample, and binding the separated target nucleic acid molecules with one or more second target endonucleases that are specific to a second locus of the target region of the target nucleic acid molecules, and uses thereof.

The present invention belongs to the fields of biomedical technology and molecular diagnosis. Disclosed is a high-throughput detection method for a rare mutation of a gene, comprising: designing specific probes; connecting Y-shaped universal linkers to a test DNA subjected to fragmentation processing, and performing amplification and enrichment of a target site by universal sequence combination of the specific probes and the linkers; performing genomic sequence alignment on sequences to be sequenced; sorting and analyzing said sequences at the same starting and ending positions, and filtering sequencing errors; and after the data filtering, the sequencing depth count of a reference allele of the target site being a, and the sequencing depth count of other alleles being b, and thus the actual mutation ratio of the site being b/(a+b). This technique can perform, by DNA fragmentation, universal linker connection, multiplex PCR amplification of specific primers and linker sequence primers, and high-throughput high-depth sequencing, enrichment and parallel sequencing on a plurality of sites to be tested.

The present invention belongs to the fields of biomedical technology and molecular diagnosis. Disclosed is a high-throughput detection method for a rare mutation of a gene, comprising: designing specific probes; connecting Y-shaped universal linkers to a test DNA subjected to fragmentation processing, and performing amplification and enrichment of a target site by universal sequence combination of the specific probes and the linkers; performing genomic sequence alignment on sequences to be sequenced; sorting and analyzing said sequences at the same starting and ending positions, and filtering sequencing errors; and after the data filtering, the sequencing depth count of a reference allele of the target site being a, and the sequencing depth count of other alleles being b, and thus the actual mutation ratio of the site being b/(a+b). This technique can perform, by DNA fragmentation, universal linker connection, multiplex PCR amplification of specific primers and linker sequence primers, and high-throughput high-depth sequencing, enrichment and parallel sequencing on a plurality of sites to be tested.

The present disclosure relates to compositions and methods for nucleic acid sequencing, and specifically, at least in certain aspects, provides methods and compositions for enhancing the efficacy, throughput and/or yield of known long-range sequencing platforms, by providing chimeric arrays of input sequences. Such arrays of component nucleic acid sequence elements can be prepared via methods that minimize introduction of bias. The application of the current methods to obtain isoform sequencing information, e.g., from patient samples is specifically also provided, as are methods for mitochondrial lineage tracing that employ the instant chimeric amplicon sequencing processes. Methods and systems for array nucleic acid sequence processing and interpretation are also provided.

The present disclosure relates to compositions and methods for nucleic acid sequencing, and specifically, at least in certain aspects, provides methods and compositions for enhancing the efficacy, throughput and/or yield of known long-range sequencing platforms, by providing chimeric arrays of input sequences. Such arrays of component nucleic acid sequence elements can be prepared via methods that minimize introduction of bias. The application of the current methods to obtain isoform sequencing information, e.g., from patient samples is specifically also provided, as are methods for mitochondrial lineage tracing that employ the instant chimeric amplicon sequencing processes. Methods and systems for array nucleic acid sequence processing and interpretation are also provided.

Methods and compositions for determining the proximity of two barcoding oligonucleotides (e.g., in a single partition or adjacent on a tissue section) using a determination of the presence of a 9 bp sequence resulting from tagmentation in different nucleic acid fragments linked to different barcoding oligonucleotides is provided.

Methods and compositions for determining the proximity of two barcoding oligonucleotides (e.g., in a single partition or adjacent on a tissue section) using a determination of the presence of a 9 bp sequence resulting from tagmentation in different nucleic acid fragments linked to different barcoding oligonucleotides is provided.

The present disclosure provides methods of processing or analyzing a sample. A method for processing a sample may comprise hybridizing a probe molecule to a target region of a nucleic acid molecule (e.g., a ribonucleic acid (RNA) molecule), barcoding the probe-nucleic acid molecule complex, and performing extension, denaturation, and amplification processes. A method for processing a sample may comprise hybridizing first and second probes to adjacent or non-adjacent target regions of a nucleic acid molecule (e.g., an RNA molecule), linking the first and second probes to provide a probe-linked nucleic acid molecule, and barcoding the probe-linked nucleic acid molecule. One or more processes of the methods described herein may be performed within a partition, such as a droplet or well. One or more processes of the methods described herein may be performed on a cell, such as a permeabilized cell.

The present disclosure provides methods of processing or analyzing a sample. A method for processing a sample may comprise hybridizing a probe molecule to a target region of a nucleic acid molecule (e.g., a ribonucleic acid (RNA) molecule), barcoding the probe-nucleic acid molecule complex, and performing extension, denaturation, and amplification processes. A method for processing a sample may comprise hybridizing first and second probes to adjacent or non-adjacent target regions of a nucleic acid molecule (e.g., an RNA molecule), linking the first and second probes to provide a probe-linked nucleic acid molecule, and barcoding the probe-linked nucleic acid molecule. One or more processes of the methods described herein may be performed within a partition, such as a droplet or well. One or more processes of the methods described herein may be performed on a cell, such as a permeabilized cell.