Methods and apparatus for nucleic acid sequencing by signal stretching and data integration
First Claim
1. The nucleic acid sequencing apparatus comprising:
- a first reaction chamber comprising non-labeled nucleotide precursors and only a single type of labeled nucleotide precursor being a first labeled nucleotide precursor, the first labeled nucleotide precursor consisting of labeled deoxyadenosine-5′
-triphosphate (dATP), a copy of a template nucleic acid, a synthetic reagent and a primer, the first reaction chamber configured to synthesize a first labeled nucleic acid strand comprising the first labeled nucleotide precursor and complementary to the template nucleic acid;
a second reaction chamber comprising non-labeled nucleotide precursors and only a single type of labeled nucleotide precursor being a second labeled nucleotide precursor, the second labeled nucleotide precursor consisting of labeled deoxycytosine-5′
-triphosphate (dCTP), a copy of the template nucleic acid, a synthetic reagent and a primer, the second reaction chamber configured to synthesize a second labeled nucleic acid strand comprising the second labeled nucleotide precursor and complementary to the template nucleic acid;
a third reaction chamber comprising non-labeled nucleotide precursors and only a single type of labeled nucleotide precursor being a third labeled nucleotide precursor, the third labeled nucleotide precursor consisting of labeled deoxythymidine-5′
-triphosphate (dTTP), a copy of the template nucleic acid, a synthetic reagent and a primer, the third reaction chamber configured to synthesize a third labeled nucleic acid strand comprising the third labeled nucleotide precursor and complementary to the template nucleic acid;
a fourth reaction chamber comprising non-labeled nucleotide precursors and only a single type of labeled nucleotide precursor being a fourth labeled nucleotide precursor, the forth labeled nucleotide precursor consisting of labeled deoxyuridine-5′
-triphosphate (dUTP), a copy of the template nucleic acid, a synthetic reagent and a primer, the fourth reaction chamber configured to synthesize a fourth labeled nucleic acid strand comprising the fourth labeled nucleotide precursor and complementary to the template nucleic acid;
one or more fluid channels operably coupled to the reaction chambers;
one or more nanopores in each of the reaction chambers, each nanopore being sized to allow the passing of the labeled nucleic acid strands through the nanopore;
a detection unit configured to detect the labeled nucleotide precursors in the labeled nucleic acid strands as the labeled nucleic acid strands pass through the nanopores.
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Abstract
The methods and apparatus 100 disclosed herein concern DNA sequencing. In some embodiments of the invention, the methods comprise measuring the distance between labeled nucleotides 220, such as nucleotides labeled with bulky groups. The methods may further comprise placing identical template DNA 200 into four reaction chambers 110, 120, 130, 140, each containing a different labeled nucleotide precursor, synthesizing complementary strands 230, 240, 250 and detecting labeled nucleotides 220. The distances between labeled nucleotides 220 may be used to construct 450 distance maps 310, 320, 330, 340 for each type of labeled nucleotide 220. The distance maps 310, 320, 330, 340 may be aligned 520 to obtain a nucleic acid sequence 210. Overlapping data analysis and frequency analysis may be used to construct 450 the distance maps 310, 320, 330, 340 and non-overlapping data analysis may be used to align 520 the distance maps into a sequence 210.
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Citations
21 Claims
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1. The nucleic acid sequencing apparatus comprising:
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a first reaction chamber comprising non-labeled nucleotide precursors and only a single type of labeled nucleotide precursor being a first labeled nucleotide precursor, the first labeled nucleotide precursor consisting of labeled deoxyadenosine-5′
-triphosphate (dATP), a copy of a template nucleic acid, a synthetic reagent and a primer, the first reaction chamber configured to synthesize a first labeled nucleic acid strand comprising the first labeled nucleotide precursor and complementary to the template nucleic acid;a second reaction chamber comprising non-labeled nucleotide precursors and only a single type of labeled nucleotide precursor being a second labeled nucleotide precursor, the second labeled nucleotide precursor consisting of labeled deoxycytosine-5′
-triphosphate (dCTP), a copy of the template nucleic acid, a synthetic reagent and a primer, the second reaction chamber configured to synthesize a second labeled nucleic acid strand comprising the second labeled nucleotide precursor and complementary to the template nucleic acid;a third reaction chamber comprising non-labeled nucleotide precursors and only a single type of labeled nucleotide precursor being a third labeled nucleotide precursor, the third labeled nucleotide precursor consisting of labeled deoxythymidine-5′
-triphosphate (dTTP), a copy of the template nucleic acid, a synthetic reagent and a primer, the third reaction chamber configured to synthesize a third labeled nucleic acid strand comprising the third labeled nucleotide precursor and complementary to the template nucleic acid;a fourth reaction chamber comprising non-labeled nucleotide precursors and only a single type of labeled nucleotide precursor being a fourth labeled nucleotide precursor, the forth labeled nucleotide precursor consisting of labeled deoxyuridine-5′
-triphosphate (dUTP), a copy of the template nucleic acid, a synthetic reagent and a primer, the fourth reaction chamber configured to synthesize a fourth labeled nucleic acid strand comprising the fourth labeled nucleotide precursor and complementary to the template nucleic acid;one or more fluid channels operably coupled to the reaction chambers; one or more nanopores in each of the reaction chambers, each nanopore being sized to allow the passing of the labeled nucleic acid strands through the nanopore; a detection unit configured to detect the labeled nucleotide precursors in the labeled nucleic acid strands as the labeled nucleic acid strands pass through the nanopores. - View Dependent Claims (2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21)
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Specification