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US 2025/0051840 A1Published application
Detection of genetic and epigenetic information in a single workflow
AssigneeFoundation Medicine, Inc.
InventorsYexun WANG, Nicole Jacinda LAMBERT, Becky WONG, Yu ZHENG, Haleigh WOOD
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Abstract
Provided herein are methods related to detecting genetic and epigenetic information in a single workflow, as well as methods of treatment, uses, systems, and computer readable storage media related thereto. These methods allow, e.g., for detection of genetic variants and epigenetic modifications (e.g., methylation level) in a single workflow and/or from a single sample (e.g., a DNA sample).
Claims41
A method of detecting genetic and epigenetic information in a single workflow, comprising:
- providing a plurality of first single-stranded DNA fragments;
- subjecting the plurality of first single-stranded DNA fragments to a round of primer extension in the presence of:
- (a) a primer that anneals to at least a portion of the first single-stranded DNA fragments, (b) a nucleic acid polymerase, and (c) a mixture of nucleotides comprising a cytosine analog that is resistant to cytosine conversion, thereby generating a plurality of first strands corresponding to the first single-stranded DNA fragments and a plurality of second strands that are complementary to the first strands, wherein the second strands comprise the cytosine analog that is resistant to cytosine conversion;
- subjecting the pluralities of the first and second strands to a cytosine conversion treatment under conditions such that unmethylated cytosine(s) if present in the first strands undergo cytosine conversion;
- and
- detecting methylation information from at least a portion of the plurality of first strands and detecting sequence information from at least a portion of the plurality of second strands.
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A method of detecting genetic and epigenetic information in a single workflow, comprising:
- providing a plurality of first single-stranded DNA fragments;
- attaching a first adaptor nucleic acid onto at least a portion of the first single-stranded DNA fragments of the plurality;
- subjecting the plurality of first single-stranded DNA fragments to a round of primer extension in the presence of:
- (a) a primer that anneals to at least a portion of the first single-stranded DNA fragments downstream or at the first adaptor nucleic acid, (b) a nucleic acid polymerase, and (c) a mixture of nucleotides comprising a cytosine analog that is resistant to cytosine conversion, thereby generating a plurality of first strands comprising the first single-stranded DNA fragments with the first adaptor nucleic acid and a plurality of second strands comprising second single-stranded DNA fragments complementary to the first single-stranded DNA fragments and a second adaptor nucleic acid complementary to the first adaptor nucleic acid, wherein the second strands comprise the cytosine analog that is resistant to cytosine conversion;
- subjecting the pluralities of the first and second strands to a cytosine conversion treatment under conditions such that unmethylated cytosine(s) if present in the first strands undergo cytosine conversion, wherein after cytosine conversion, the sequences of the first and second adaptor nucleic acids are no longer complementary;
- and
- detecting methylation information from at least a portion of the plurality of first strands and detecting sequence information from at least a portion of the plurality of second strands.
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A method of detecting genetic and epigenetic information in a single workflow, comprising:
- providing a plurality of first single-stranded DNA fragments;
- subjecting the plurality of first single-stranded DNA fragments to a round of primer extension in the presence of:
- (a) a primer comprising:
- (i) a second adaptor nucleic acid portion that does not anneal to the first single-stranded DNA fragments and (ii) a portion that anneals to at least a portion of the first single-stranded DNA fragments, (b) a nucleic acid polymerase, and (c) a mixture of nucleotides comprising a cytosine analog that is resistant to cytosine conversion, thereby generating a plurality of first strands comprising the first single-stranded DNA fragments with a first adaptor nucleic acid complementary to the second adaptor nucleic acid and a plurality of second strands comprising second single-stranded DNA fragments complementary to the first single-stranded DNA fragments and the second adaptor nucleic acid non-complementary to the first adaptor nucleic acid, wherein the second strands comprise the cytosine analog that is resistant to cytosine conversion;
- subjecting the pluralities of the first and second strands to a cytosine conversion treatment under conditions such that unmethylated cytosine(s) if present in the first strands undergo cytosine conversion, wherein after cytosine conversion, the sequences of the first and second adaptor nucleic acids are no longer complementary;
- and
- detecting methylation information from at least a portion of the plurality of first strands and detecting sequence information from at least a portion of the plurality of second strands.
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Dependencies are inferred from the claim text.