Geometric synthesis methods and compositions for double-stranded nucleic acid sequencing
AssigneeCamena Bioscience Limited
InventorNeil BELL
Life of this patent
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Abstract
The present disclosure provides compositions, kits and methods for sequencing double-stranded nucleic acids. The compositions, kits and methods comprise partially double-stranded identifier molecules and partially double-stranded adapter molecules. The compositions, kits and methods can be used to determine the abundance and/or identity of specific transcripts in a plurality of double-stranded nucleic acids, as well as identifying the frequency of mutations within certain transcripts in the plurality of double-stranded nucleic acids.
Claims26
A plurality of partially double-stranded identifier molecules,
- wherein the partially double-stranded identifier molecules comprise:
- a double-stranded region comprising an identifier sequence;
- and
- a first overhang;
- wherein the plurality comprises at least about 12 species of the partially double-stranded adapter molecules, wherein each species of partially double-stranded adapter molecules comprises an identifier sequence that is different from the identifier sequence of any other species of partially double-stranded adapter molecules in the plurality,
- wherein the identifier sequence of one species of partially double-stranded identifier molecules will have a hamming distance of at least about two to any other identifier sequence of any other species of partially double-stranded identifier molecules in the plurality.
The plurality of partially double-stranded identifier molecules of any of the preceding claims, wherein the first and/or second overhangs are
- a) 5′ overhangs;
- or
- b) 3′ overhangs.
The plurality of partially double-stranded identifier molecules of any of the preceding claims, wherein:
- a) the identifier sequence spans the entire double-stranded region;
- or
- b) the identifier sequence spans a portion of the double-stranded region.
The plurality of partially double-stranded identifier molecules of any of the preceding claims, wherein the identifier sequence is:
- a) about 9 nucleotides in length;
- b) about 10 nucleotides in length:
- c) about 11 nucleotides in length;
- d) about 12 nucleotides in length;
- e) about 19 nucleotides in length;
- f) about 20 nucleotides in length;
- g) about 21 nucleotides in length;
- or
- h) about 22 nucleotides in length.
The plurality of partially double-stranded identifier molecules of any of the preceding claims, wherein the first overhang and/or the second overhang is about 1 nucleotide in length, preferably wherein the first overhang and/or the second overhang is:
- a) an adenine or a thymine;
- or
- b) a guanosine or a cytosine.
The plurality of partially double-stranded identifier molecules of any of the preceding claims, wherein the first overhang and/or the second overhang is:
- a) about 2 nucleotides in length;
- b) about 3 nucleotides in length;
- c) about 4 nucleotides in length;
- or
- d) about 5 nucleotides in length.
The plurality of partially double-stranded identifier molecules of any of the preceding claims, wherein the partially double-stranded identifier molecules comprise DNA.
The plurality of partially double-stranded identifier molecules of any of the preceding claims, wherein the plurality comprises:
- a) at least about 24 species of the partially double-stranded identifier molecules;
- b) at least about 48 species of the partially double-stranded identifier molecules;
- or
- c) at least about 96 species of the partially double-stranded identifier molecules.
A plurality of partially double-stranded adapter molecules,
- wherein the partially double-stranded adapter molecules comprise:
- a double-stranded region;
- an overhang;
- a single-stranded 5′ arm;
- and
- a single-stranded 3′ arm;
- wherein the single-stranded 5′ arm comprises at least one amplification primer binding site and the single-stranded 3′ arm comprises at least one amplification primer binding site.
Dependencies are inferred from the claim text.