IP Library › Granted Patent US 12,460,261
Granted Patent B2
US 12,460,261 · App. 18/988,274 · Granted Nov 4, 2025

Compositions and methods for nucleic acid sequencing

Inventors: Eli N. Glezer (Del Mar, CA); Mohammad Vatankhah Varnosfaderani (San Marcos, CA); Daan Witters (San Diego, CA); Vahid Karimkhani (San Diego, CA)
Assignee: Singular Genomics Systems, Inc.
C12Q1/6869C12Q1/6844C12Q1/6874G01N33/5308G01N33/544C12Q2533/101C12Q2565/601
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Quick Facts
Patent No.
US 12,460,261
App. No.
18/988,274
Filed
Dec 19, 2024
Granted
Nov 4, 2025
Kind
B2
Art Unit
1683
USPC
435/6.11
Abstract

Provided herein are methods and compositions for improved sequencing techniques using, for example, polymeric particles and/or three-dimensional structures.

Claims (23)

1. A method of detecting polynucleotides in a polymer scaffold, the method comprising:

(a) amplifying polynucleotides to produce discrete amplicon clusters within a three-dimensional polymer scaffold, wherein the amplicons are covalently bound to the polymer scaffold;

(b) contacting the polymer scaffold with a plurality of fluorescently labeled molecules and, within two or more discrete amplicon clusters, binding a fluorescently labeled molecule to a nucleic acid molecule in the amplicon cluster;

(c) detecting the fluorescently labeled molecules to identify the polynucleotide in the polymer scaffold.

2. The method of claim 1 , wherein detecting the fluorescently labeled molecules further identifies three-dimensional coordinates of the polynucleotide.

3. The method of claim 2 , further comprising forming an image based on the detected labels.

4. The method of claim 1 , prior to (a), comprising contacting the polymer scaffold with a plurality of primers and covalently attaching the primers to the polymer scaffold.

5. The method of claim 4 , wherein the plurality comprises a first primer comprising a first capture sequence and a second primer comprising a second capture sequence.

6. The method of claim 5 , comprising binding a first template polynucleotide to the first capture sequence, wherein the first template polynucleotide comprises a first primer binding sequence.

7. The method of claim 1 , comprising cross-linking the amplicons to the polymer scaffold.

8. The method of claim 1 , wherein the three-dimensional polymer scaffold is in the channel of a flow cell.

9. The method of claim 1 , wherein amplifying comprises isothermal amplification.

10. The method of claim 1 , wherein amplifying comprises rolling circle amplification.

11. The method of claim 1 , wherein the fluorescently labeled molecules are fluorescently labeled nucleotides.

12. The method of claim 11 , wherein each fluorescently labeled nucleotide comprises a fluorophore moiety covalently attached to a nucleotide molecule via a cleavable linker.

13. The method of claim 12 , further comprising cleaving the cleavable linker and removing the fluorophore moiety and repeating (b) and (c).

14. The method of claim 13 , wherein detecting comprises imaging multiple two-dimensional planes.

15. The method of claim 1 , wherein the fluorescently labeled molecules are fluorescently labeled oligonucleotides.

16. The method of claim 15 , further comprising removing the fluorescently labeled oligonucleotides and repeating (b) and (c).

17. The method of claim 16 , wherein detecting comprises imaging multiple two-dimensional planes.

18. The method of claim 1 , wherein the polynucleotides comprise a gene sequence or a portion thereof.

19. The method of claim 1 , wherein the polymer scaffold comprises polyacrylamide.

20. The method of claim 1 , wherein the polymer scaffold comprises polyacrylate.

Assignments (2)
ASSIGNMENT OF ASSIGNOR'S INTEREST Recorded Apr 18, 2025
From: GLEZER, ELI N.; VARNOSFADERANI, MOHAMMAD VATANKHAH; WITTERS, DAAN; KARIMKHANI, VAHID
To: SINGULAR GENOMICS SYSTEMS, INC.
Reel/Frame 070886/0530 →
ASSIGNMENT OF ASSIGNOR'S INTEREST Recorded Feb 18, 2025
From: GLEZER, ELI N.; VARNOSFADERANI, MOHAMMAD VATANKHAH; WITTERS, DAAN; KARIMKHANI, VAHID
To: SINGULAR GENOMICS SYSTEMS, INC.
Reel/Frame 070246/0564 →
Continuity (5)
Continuation 17542343 · Dec 3, 2021
Continuation 17030886 · Sep 24, 2020
Continuation PCTUS2020017060 · Feb 6, 2020
Provisional Application 62802062 · Feb 6, 2019
Related Publication 20250122560A1 · Apr 17, 2025
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