Solution-based methods and materials for sequence analysis by hybridization
First Claim
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1. A method of detecting a sequence of a target nucleic acid, comprising:
- (a) contacting a target nucleic acid with one or more mixtures of a plurality of oligonucleotide probe molecules of predetermined length and predetermined sequence, wherein each probe molecule comprises an information region and at least two probe molecules have different information regions, under conditions which produce, on average, more probe;
target hybridization with probe molecules which are perfectly complementary to the target nucleic acid in the information region of the probe molecules than with probe molecules which are mismatched in the information region, wherein the target nucleic acid is not attached to a support, and wherein the probe molecules are not attached to a support;
(b) detecting probe molecules that hybridize with the target nucleic acid, using a reader capable of detecting an individual probe molecule; and
(c) detecting a sequence of the target nucleic acid by overlapping sequences of the information regions of at least two of the probe molecules contacted with the target in step (a).
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Abstract
Novel solution-based methods and materials, including apparatus, for sequence analysis by hybridization are provided.
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Citations
48 Claims
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1. A method of detecting a sequence of a target nucleic acid, comprising:
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(a) contacting a target nucleic acid with one or more mixtures of a plurality of oligonucleotide probe molecules of predetermined length and predetermined sequence, wherein each probe molecule comprises an information region and at least two probe molecules have different information regions, under conditions which produce, on average, more probe;
target hybridization with probe molecules which are perfectly complementary to the target nucleic acid in the information region of the probe molecules than with probe molecules which are mismatched in the information region, wherein the target nucleic acid is not attached to a support, and wherein the probe molecules are not attached to a support;
(b) detecting probe molecules that hybridize with the target nucleic acid, using a reader capable of detecting an individual probe molecule; and
(c) detecting a sequence of the target nucleic acid by overlapping sequences of the information regions of at least two of the probe molecules contacted with the target in step (a). - View Dependent Claims (2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 34, 35, 36, 37)
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31. A method of sequencing a target nucleic acid, comprising:
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(a) contacting a target nucleic acid with one or more mixtures of a plurality of oligonucleotide probe molecules of predetermined length and predetermined sequence, wherein each probe molecule comprises an information region and at least two probe molecules have different information regions, under conditions which produce, on average, more probe;
target hybridization with probe molecules which are perfectly complementary to the target nucleic acid in the information region of the probe molecules than with probe molecules which are mismatched in the information region, wherein the target nucleic acid is not attached to a support, and wherein the probe molecules are not attached to a support;
(b) covalently joining probe molecules that form contiguous probe;
target hybrids that are perfectly complementary to the target in the information region of the probe molecules; and
(c) detecting covalently joined probe molecules, using a reader capable of detecting an individual probe molecule. - View Dependent Claims (32, 33)
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- 38. A kit comprising a mixture of probe molecules, wherein about 100 or more probe molecules each have distinct information regions, wherein two or more of the sequences of said distinct information regions within the mixture overlap
- 43. A kit comprising a set of mixtures of probe molecules, wherein about 100 or more probe molecules each have distinct information regions, wherein two or more of the sequences of said distinct information regions within the set overlap
- 47. A tag which is a bar code comprising an alternating arrangement of elements of varying detectable properties, wherein consecutive elements have a difference in at least one of their detectable properties.
Specification