Compositions and methods for sequencing nucleic acids
AssigneeSeqWell, Inc.
InventorsJoseph C. MELLOR, Jack T. LEONARD
Life of this patent
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Abstract
The invention provides compositions, including tethered synaptic complexes (TSCs), artificial nucleic acids, molecular constructs that include artificial nucleic acids bound to transposases, and kits; as well as methods of using the same, for example, for preparation of nucleic acid libraries and sequencing.
Claims129
A tethered synaptic complex (TSC) comprising:
- a first artificial nucleic acid comprising a first end comprising a first transposase binding site (TBS), a second end comprising a second TBS, and a linking segment disposed between the first TBS and the second TBS;
- a second artificial nucleic acid comprising a first end comprising a first TBS;
- a third artificial nucleic acid comprising a first end comprising a first TBS;
- a first synaptic complex comprising a first pair of oligomerized transposases, the first pair comprising a first transposase and a second transposase, wherein the first transposase is bound to the first TBS of the first artificial nucleic acid, and the second transposase is bound to the first TBS of the second artificial nucleic acid;
- and
- a second synaptic complex comprising a second pair of oligomerized transposases, the second pair comprising a third transposase and a fourth transposase, wherein the third transposase is bound to the second TBS of the first artificial nucleic acid, and the fourth transposase is bound to the first TBS of the third artificial nucleic acid.
An artificial nucleic acid comprising a first end comprising a first transposase binding site (TBS), a second end comprising a second TBS, and a linking segment disposed between the first TBS and the second TBS, wherein upon binding of a first transposase to the first TBS and a second transposase to the second TBS, the first transposase does not oligomerize with the second transposase.
A method of preparing a TSC, the method comprising:
- (a) providing at least a first synaptic complex and a second synaptic complex, wherein
- (i) the first synaptic complex comprises a first pair of oligomerized transposases, wherein one member of the pair is bound to a first nucleic acid comprising a TBS and a sticky end;
- and
- (ii) the second synaptic complex comprises a second pair of oligomerized transposases, wherein one member of the pair is bound to a second nucleic acid comprising a TBS and a sticky end;
- and
- (b) ligating the first synaptic complex to the second synaptic complex by ligating the sticky ends of the first nucleic acid and the second nucleic acid in a ligation reaction,
- thereby preparing a TSC.
A synaptic complex comprising:
- (a) a first transposase;
- (b) a second transposase;
- (c) a first artificial nucleic acid comprising a first end comprising a TBS and being linked to a first conjugating moiety other than a 3′ hydroxyl;
- and
- (d) a second artificial nucleic acid comprising a first end comprising a TBS and being linked to a second conjugating moiety other than a 3′ hydroxyl,
- wherein the first transposase is bound to the TBS of the first artificial nucleic acid, the second transposase is bound to the TBS of the second artificial nucleic acid, and the first transposase and the second transposase are oligomerized.
A method of preparing a TSC, the method comprising:
- (a) providing at least a first synaptic complex and a second synaptic complex, wherein
- (i) the first synaptic complex comprises a first pair of oligomerized transposases, wherein one member of the pair is bound to a first nucleic acid comprising a TBS and being linked to a first component of an affinity binding pair or a first conjugating moiety;
- and
- (ii) the second synaptic complex comprises a second pair of oligomerized transposases, wherein one member of the pair is bound to a second nucleic acid comprising a TBS and being linked to a second component of the affinity binding pair or a second conjugating moiety;
- and
- (b) linking the first synaptic complex to the second synaptic complex, wherein linking of the first conjugating moiety to the second conjugating moiety does not result in a phosphodiester bond,
- thereby preparing a TSC.
Dependencies are inferred from the claim text.