Systems and methods for sequencing by hybridization
First Claim
1. A gapped probe, comprising a sequence of universal and designate nucleotides and/or nucleotide analogs ordered in a predetermined pattern, wherein the pattern has a reduced shift-overlap count.
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Abstract
The systems and methods described herein relate to nucleic acid probes comprising a pattern of universal and designate nucleotides (or analogs thereof), or ‘gapped’ probes, and the use of sets of gapped probes in sequencing by hybridization to determine the sequence of nucleic acid sequences. The inclusion of universal nucleotides in the probes allows for efficient and rapid sequencing of longer nucleotide sequences than can be sequenced using traditional probes. The systems and methods described herein also relate to apparatus for sequencing nucleic acids which include gapped probes, as well as computer systems capable of analyzing data generated using gapped probes in such apparatus.
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Citations
39 Claims
- 1. A gapped probe, comprising a sequence of universal and designate nucleotides and/or nucleotide analogs ordered in a predetermined pattern, wherein the pattern has a reduced shift-overlap count.
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6. A gapped probe, comprising a sequence of universal and designate nucleotides and/or nucleotide analogs ordered in a pattern, wherein
(a) the pattern comprises a first string of universal nucleotides followed by a first segment, and a second string of universal nucleotides followed by a second segment, (b) the first string and the second string each comprise two or more consecutive universal nucleotides; -
(c) the first segment and the second segment each comprise a designate nucleotide; and
wherein at least one of the designate nucleotides is a nucleotide analog selected from the group consisting of;
a peptide nucleic acid, pyranosyl-RNA, a hexitol nucleic acid, a mannitol nucleic acid, an altritol nucleic acid, a 2′
-5′
nucleic acid, a locked nucleic acid, a seco-locked nucleic acid, a bicyclo-DNA, a tricyclo-DNA, 3-hydroxy-N-acetylprolinol substituted nucleic acid, a carbocyclic nucleic acid, a carbocyclic/bicyclic nucleic acid, a nucleic acid with a triazole backbone, a nucleic acid with an imidazole backbone, a 1-phenylserinol nucleic acid, a nucleic acid with an alpha anomer backbone, and a metal-linked nucleic acid.
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7. An gapped probe, comprising a sequence of universal and designate nucleotides and/or nucleotide analogs ordered in a pattern, wherein the pattern comprises a root and an iterated unit, and wherein the length of the root is identical to the length of the iterated unit, and wherein the probe comprises at least one nucleotide analog selected from the group consisting of:
- a peptide nucleic acid, pyranosyl-RNA, a hexitol nucleic acid, a mannitol nucleic acid, an altritol nucleic acid, a 2′
-5′
nucleic acid, a locked nucleic acid, a seco-locked nucleic acid, a bicyclo-DNA, a bicyclo[3.3.0]DNA, a tricyclo-DNA, 3-hydroxy-N-acetylprolinol substituted nucleic acid, a carbocyclic nucleic acid, a carbocyclic/bicyclic nucleic acid, a nucleic acid with a triazole backbone, a nucleic acid with an imidazole backbone, a 1-phenylserinol nucleic acid, a nucleic acid with an alpha anomer backbone, and a metal-linked nucleic acid. - View Dependent Claims (8)
- a peptide nucleic acid, pyranosyl-RNA, a hexitol nucleic acid, a mannitol nucleic acid, an altritol nucleic acid, a 2′
- 9. A set of gapped probes, comprising a plurality of instances of a sequence of universal and designate nucleotides and/or nucleotide analogs ordered in a predetermined pattern, wherein the pattern has a reduced shift-overlap count.
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17. A probe array, comprising
a substrate, and a set of gapped probes disposed thereon, wherein each probe comprises an instance of a predetermined pattern of universal and designate nucleotides and/or nucleotide analogs such that the set comprises a plurality of instances of the pattern.
- 25. A set of gapped probes, comprising a plurality of instances of a sequence of universal and designate nucleotides and/or nucleotide analogs ordered in a predetermined pattern, wherein the each gapped probe of the set comprises greater than five nucleotides and/or nucleotide analogs.
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27. A method for analyzing a nucleic acid sequence, comprising
providing a set of probes wherein each probe comprises an instance of a pattern of universal and designate nucleotides and/or nucleotide analogs such that the set comprises a plurality of instances of the pattern, determining a spectrum of probes representative of the probes in the set of probes which hybridize to a test sequence, and ordering the spectrum of probes to determine a sequence of a portion of the test sequence.
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28. A method for ordering a spectrum of probes to determine a sequence of a portion of a test sequence, comprising
i) providing a spectrum of probes that hybridize to a test sequence, wherein each probe in the spectrum is an instance of a pattern of universal and designate nucleotides and/or nucleotide analogs, which pattern requires a designate nucleotide and/or nucleotide analog at an mth position and an nth position, ii) identifying a first subset of probes from the spectrum whose first m− - 1 nucleotides and/or nucleotide analogs correspond to a last m−
1 nucleotides of a growing sequence,iii) appending the nucleotide at the mth position to the growing sequence if a single nucleotide and/or nucleotide analog occurs at the mth position of all probes in the first subset. - View Dependent Claims (29, 30, 31, 37, 39)
- 1 nucleotides and/or nucleotide analogs correspond to a last m−
Specification