Sequencing by Structure Assembly
AssigneePresident and Fellows of Harvard College
InventorsGeorge M. Church, Richard C. Terry, Frederic Vigneault, Francois Vigneault
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Abstract
A method of sequencing nucleic acids is provided using sequencing by ligation and/or sequencing by hybridization.
Claims36
A method for determining the sequence of nucleotides in a target polynucleotide comprising
- (a) hybridizing a sequencing primer to a single stranded nucleic acid template,
- (b) hybridizing an oligonucleotide probe to the single stranded nucleic acid template and ligating the oligonucleotide probe to the sequencing primer to form an extended hybridized sequence, wherein the oligonucleotide probe includes a template-nonhybridizing nucleic acid structure cleavably attached thereto and wherein the template-nonhybridizing nucleic acid structure has a cleavable nucleotide immediately attached to a terminal hybridized nucleotide of the oligonucleotide probe,
- (d) identifying a nucleotide in the oligonucleotide probe and identifying a complementary nucleotide in the single stranded nucleic acid, and optionally
- (e) removing the template-nonhybridizing nucleic acid structure by cleavage of the cleavable nucleotide and generating an extendable terminus on the extended hybridized sequence.
A method for determining the sequence of nucleotides in a target polynucleotide comprising
- (a) hybridizing a sequencing primer to a single stranded nucleic acid template,
- (b) hybridizing a dual probe structure including a first oligonucleotide probe, a template non-hybridizing nucleic acid structure and a second oligonucleotide probe to the single stranded nucleic acid template and ligating the first oligonucleotide probe to the sequencing primer to form an extended hybridized sequence, and
- (c) identifying a nucleotide in the first or second oligonucleotide probe and identifying a complementary nucleotide in the single stranded nucleic acid.
An oligonucleotide probe comprising a template-hybridizing nucleic acid structure and a template-non-hybridizing nucleic acid structure attached thereto by a cleavable nucleotide immediately attached to a terminal hybridizable nucleotide of the oligonucleotide probe.
A dual oligonucleotide probe comprising a first oligonucleotide probe, a template non-hybridizing nucleic acid structure and a second oligonucleotide probe.
Dependencies are inferred from the claim text.