The invention relates to methods for pairwise sequencing of a double-stranded polynucleotide template, which permit the sequential determination of nucleotide sequences in two distinct and separate regions on complementary strands of the double-stranded polynucleotide template. The two regions for sequence determination may or may not be complementary to each other.

The invention relates to methods for pairwise sequencing of a double-stranded polynucleotide template, which permit the sequential determination of nucleotide sequences in two distinct and separate regions on complementary strands of the double-stranded polynucleotide template. The two regions for sequence determination may or may not be complementary to each other.

DNA damage (e.g., cytosine deamination) can appear more frequently in hypermethylated partitions of DNA (e.g., cell-free DNA) samples, than in hypomethylated partitions. Embodiments include sequencing hypermethylated partitions and hypomethylated partitions wherein calling a C to T or G to A transition mutation relative to a reference sequence based on sequences of molecules from the hypermethylated partition requires observation of the transition mutation in a greater number of molecules than calling a C to T or G to A transition mutation relative to the reference sequence based on sequences of molecules from the hypomethylated partition, or C to T or G to A transition mutations are not called relative to a reference sequence based on sequences of molecules of the hypermethylated partition.

DNA damage (e.g., cytosine deamination) can appear more frequently in hypermethylated partitions of DNA (e.g., cell-free DNA) samples, than in hypomethylated partitions. Embodiments include sequencing hypermethylated partitions and hypomethylated partitions wherein calling a C to T or G to A transition mutation relative to a reference sequence based on sequences of molecules from the hypermethylated partition requires observation of the transition mutation in a greater number of molecules than calling a C to T or G to A transition mutation relative to the reference sequence based on sequences of molecules from the hypomethylated partition, or C to T or G to A transition mutations are not called relative to a reference sequence based on sequences of molecules of the hypermethylated partition.

The present technology relates generally to methods and compositions for targeted nucleic acid sequence enrichment, as well as uses of such enrichment for error-corrected nucleic acid sequencing applications. In some embodiments, highly accurate, error corrected and massively parallel sequencing of nucleic acid material is possible using a combination of uniquely labeled strands in a double-stranded nucleic acid complex in such a way that each strand can be informatically related to its complementary strand, but also distinguished from it following sequencing of each strand or an amplified product derived therefrom. In various embodiments, this information can be used for the purpose of error correction of the determined sequence.

The present technology relates generally to methods and compositions for targeted nucleic acid sequence enrichment, as well as uses of such enrichment for error-corrected nucleic acid sequencing applications. In some embodiments, highly accurate, error corrected and massively parallel sequencing of nucleic acid material is possible using a combination of uniquely labeled strands in a double-stranded nucleic acid complex in such a way that each strand can be informatically related to its complementary strand, but also distinguished from it following sequencing of each strand or an amplified product derived therefrom. In various embodiments, this information can be used for the purpose of error correction of the determined sequence.

The present disclosure is concerned with compositions and methods for reducing the steps used in the generation of monoclonal clusters by combining the enzymes used for linearization and removal of unused surface primers.

The present disclosure is concerned with compositions and methods for reducing the steps used in the generation of monoclonal clusters by combining the enzymes used for linearization and removal of unused surface primers.

Aspects of the invention include methods for preparing sequencing libraries, performing sequencing procedures that can correct for process-related errors, and identifying rare variants that are or may be indicative of cancer.

Aspects of the invention include methods for preparing sequencing libraries, performing sequencing procedures that can correct for process-related errors, and identifying rare variants that are or may be indicative of cancer.