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US 2021/0340621 A1Published application
Methods and systems for sequencing
AssigneeReadcoor, LLC
InventorsEvan Daugharthy, Richard Terry
Life of this patent
Estimating the term…
Abstract
Provided herein are compositions, methods, and systems for amplifying and identifying nucleic acids within a biological sample. The compositions, methods, and systems are generally compatible with volumetric imaging techniques and samples comprising nucleic acids contained within a three-dimensional matrix.
Claims21
-37. (canceled)
A method for identifying one or more nucleotides of a nucleic acid molecule, comprising:
- (a) providing said nucleic acid molecule having hybridized thereto a sequencing primer and at least one nucleic acid probe, wherein said at least one nucleic acid probe comprises (i) a template hybridizing sequence that is complementary to a sequence of said nucleic acid molecule and (ii) a template nonhybridizing sequence, which template nonhybridizing sequence corresponds to said one or more nucleotides of said nucleic acid molecule, and wherein said template nonhybridizing sequence comprises a nucleic acid initiator;
- (b) ligating said sequencing primer to said at least one nucleic acid probe;
- (c) contacting said nucleic acid molecule with a plurality of nucleic acid amplifiers such that said nucleic acid initiator of said template nonhybridizing sequence initiates an amplification reaction of at least a subset of said plurality of nucleic acid amplifiers to form an amplification product attached to said nucleic acid initiator;
- (d) detecting a signal from said amplification product to identify one or more other nucleotides of said template nonhybridizing sequence, which one or more other nucleotides corresponds to said one or more nucleotides of said nucleic acid molecule;
- and
- (e) using at least said one or more other nucleotides identified in (d) to identify said one or more nucleotides of said nucleic acid molecule.
3 more elements
A composition comprising:
- a nucleic acid molecule having hybridized thereto a sequencing primer and at least one nucleic acid probe hybridized thereto, wherein said at least one nucleic acid probe comprises (i) a template hybridizing sequence that is complementary to a sequence of said nucleic acid molecule and (ii) a template nonhybridizing sequence, which template nonhybridizing sequence corresponds to one or more nucleotides of said nucleic acid molecule, wherein said template nonhybridizing sequence comprises a nucleic acid initiator, and wherein said sequencing primer is ligated to said at least one nucleic acid probe;
- and
- a plurality of nucleic acid amplifiers, wherein at least a subset of said plurality of nucleic acid amplifiers is configured to form an amplification product attached to said nucleic acid initiator.
A kit for identifying one or more nucleotides of a nucleic acid molecule, comprising:
- a sequencing primer;
- at least one nucleic acid probe, wherein said at least one nucleic acid probe comprises (i) a template hybridizing sequence that is complementary to a sequence of said nucleic acid molecule and (ii) a template nonhybridizing sequence, which template nonhybridizing sequence corresponds to said one or more nucleotides of said nucleic acid molecule, and wherein said template nonhybridizing sequence comprises a nucleic acid initiator;
- a plurality of nucleic acid amplifiers;
- and
- instructions that direct a user to:
- (a) provide said nucleic acid molecule having hybridized thereto said sequencing primer and said at least one nucleic acid probe;
- (b) ligate said sequencing primer to said at least one nucleic acid probe;
- (c) contact said nucleic acid molecule with said plurality of nucleic acid amplifiers such that said nucleic acid initiator of said template nonhybridizing sequence initiates an amplification reaction of at least a subset of said plurality of nucleic acid amplifiers to form an amplification product attached to said nucleic acid initiator;
- (d) detect a signal from said amplification product to identify one or more other nucleotides of said template nonhybridizing sequence, which one or more other nucleotides corresponds to said one or more nucleotides of said nucleic acid molecule;
- and
- (e) use at least said one or more other nucleotides identified in (d) to identify said one or more nucleotides of said nucleic acid molecule.
8 more elements
Dependencies are inferred from the claim text.