Capsule array devices and methods of use

Patent metadata

Field	Value
Publication number	US-10053723-B2
Application number	US-201715719459-A
Country	US
Kind code	B2
Filing date	Sep 28, 2017
Priority date	Aug 14, 2012
Publication date	Aug 21, 2018
Grant date	Aug 21, 2018

Abstract

This disclosure provides microwell capsule array devices. The microwell capsule array devices are generally capable of performing one or more sample preparation operations. Such sample preparation operations may be used as a prelude to one more or more analysis operations. For example, a device of this disclosure can achieve physical partitioning and discrete mixing of samples with unique molecular identifiers within a single unit in preparation for various analysis operations. The device may be useful in a variety of applications and most notably nucleic-acid-based sequencing, detection and quantification of gene expression and single-cell analysis.

First claim

What is claimed is:

1. A composition for use in barcoding nucleic acid molecules, comprising:
a population of at least 1,000 gel beads comprising a plurality of nucleic acid barcode molecules,
wherein a given gel bead of said at least 1,000 gel beads comprises at least 1,000 nucleic acid barcode molecules of said plurality of nucleic acid barcode molecules,
wherein at least (i) a first subset of said at least 1,000 nucleic acid barcode molecules is coupled to an interior surface of said given gel bead, and (ii) a second subset of said at least 1,000nucleic acid barcode molecules is coupled to an outer surface of said given gel bead, and
wherein each of said at least 1,000 nucleic acid barcode molecules comprises an identical barcode sequence that differs from barcode sequences of barcode molecules comprised in other gel beads of said at least 1,000 gel beads.

2. The composition of claim 1 , wherein said at least 1,000 gel beads are at least 100,000 gel beads.

3. The composition of claim 1 , wherein said given gel bead is coupled to at least 100,000 of said plurality of nucleic acid barcode molecules, wherein each of said at least 100,000 nucleic acid barcode molecules comprises said identical barcode sequence.

4. The composition of claim 1 , wherein said given gel bead is coupled to at least 1,000,000 of said plurality of nucleic acid barcode molecules, wherein each of said at least 1,000,000 nucleic acid barcode molecules comprises said identical barcode sequence.

5. The composition of claim 1 , wherein said first subset of said at least 1,000 nucleic acid barcode molecules is covalently bonded to said interior surface of said given gel bead.

6. The composition of claim 1 , wherein said first subset of said at least 1,000 nucleic acid barcode molecules is reversibly coupled to said interior surface of said given gel bead.

7. The composition of claim 6 , wherein said first subset of said at least 1,000 nucleic acid barcode molecules is reversibly coupled to said interior surface of said given gel bead through chemical cross-linkers.

8. The composition of claim 7 , wherein said chemical cross-linkers are degradable by an acid or a reducing agent.

9. The composition of claim 6 , wherein said first subset of said at least 1,000 nucleic acid barcode molecules is reversibly coupled to said interior surface of said given gel bead through disulfide bonds.

10. The composition of claim 1 , wherein said plurality of nucleic acid barcode molecules is releasable from said at least 1,000 gel beads.

11. The composition of claim 1 , wherein said at least 1,000 gel beads have substantially uniform cross-sectional sizes.

12. The composition of claim 1 , wherein said given gel bead has a cross-sectional dimension of at least 10 micrometers.

13. The composition of claim 1 , wherein said at least 1,000 gel beads are degradable upon application of a stimulus.

14. The composition of claim 1 , wherein said at least 1,000 gel beads are at least 1,000,000 gel beads.

15. The composition of claim 1 , wherein said at least 1,000 nucleic acid barcode molecules comprise immobilization sequence regions.

16. The composition of claim 1 , wherein said given gel bead comprises cross-linked polymeric precursors.

17. A composition for use in barcoding a nucleic acid molecule, comprising:
at least 1,000 nucleic acid barcode molecules coupled to a gel bead, wherein said gel bead is degradable upon application of a stimulus,
wherein at least a subset of said at least 1,000 nucleic acid barcode molecules is coupled to an interior surface of said gel bead, and
wherein each of said at least 1,000 nucleic acid barcode molecules coupled to said gel bead comprises an identical barcode sequence.

18. The composition of claim 1 , wherein said given gel bead is coupled to at least 10,000,000 of said plurality of nucleic acid barcode molecules.

19. The composition of claim 17 , wherein said gel bead comprises cross-linked polymeric precursors.

20. The composition of claim 17 , wherein said gel bead is coupled to at least 100,000 nucleic acid barcode molecules, wherein each of said at least 100,000 nucleic acid barcode molecules comprises said identical barcode sequence.

21. The composition of claim 17 , wherein said at least a subset of said at least 1,000 nucleic acid barcode molecules is covalently bonded to said interior surface of said gel bead.

22. The composition of claim 17 , wherein said at least a subset of said at least 1,000 nucleic acid barcode molecules is reversibly coupled to said interior surface of said gel bead.

23. The composition of claim 22 , wherein said at least a subset of said at least 1,000 nucleic acid barcode molecules is reversibly coupled to said interior surface of said gel bead through chemical cross-linkers.

24. The composition of claim 23 , wherein said chemical cross-linkers are degradable by an acid.

25. The composition of claim 22 , wherein said at least a subset of said at least 1,000 nucleic acid barcode molecules is reversibly coupled to said interior of said gel bead through disulfide bonds.

26. The composition of claim 23 , wherein said chemical cross-linkers are degradable by a reducing agent.

27. The composition of claim 17 , wherein an additional subset of said at least 1,000 nucleic acid barcode molecules is coupled to an outer surface of said gel bead.

28. The composition of claim 17 , wherein said at least 1,000 nucleic acid barcode molecules are releasable from said gel bead.

29. The composition of claim 17 , wherein said at least 1,000 nucleic acid barcode molecules comprise immobilization sequence regions.

30. The composition of claim 17 , wherein said at least 1,000 nucleic acid barcode molecules is at least 10,000,000 nucleic acid barcode molecules coupled to said gel bead.

Assignees

• 10X Genomics Inc

Inventors

• Hindson Benjamin
• Saxonov Serge
• Schnall-Levin Michael

Classifications

• B01L2200/0647

  Handling flowable solids, e.g. microscopic beads, cells, particles · CPC title

• C12Q2535/122

  Massive parallel sequencing · CPC title

• B01L3/508

  Rigid containers without fluid transport within · CPC title

• B01L3/523

  with means for closing or opening · CPC title

• C12Q1/6806 Primary

  Preparing nucleic acids for analysis, e.g. for polymerase chain reaction [PCR] assay (C12Q1/6804 takes precedence) · CPC title