Capsule array devices and methods of use

Patent metadata

Field	Value
Publication number	US-12037634-B2
Application number	US-202117353202-A
Country	US
Kind code	B2
Filing date	Jun 21, 2021
Priority date	Aug 14, 2012
Publication date	Jul 16, 2024
Grant date	Jul 16, 2024

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 system, comprising:
(i) a bead comprising at least 1,000,000 oligonucleotide barcodes; and
(ii) a microfluidic device comprising:
(a) a first junction of channels configured to form an aqueous mixture comprising the bead comprising the at least 1,000,000 oligonucleotide barcodes and a sample comprising a nucleic acid analyte; and
(b) a second junction of channels configured to contact the aqueous mixture with an immiscible continuous phase, thereby generating a droplet comprising (I) the bead comprising the at least 1,000,000 oligonucleotide barcodes and (II) the nucleic acid analyte.

2. The system of claim 1 , wherein oligonucleotide barcodes of the at least 1,000,000 oligonucleotide barcodes comprise a common barcode sequence.

3. The system of claim 1 , wherein oligonucleotide barcodes of the at least 1,000,000 oligonucleotide barcodes comprise a primer sequence.

4. The system of claim 3 , wherein the primer sequence is a sequencing primer sequence.

5. The system of claim 3 , wherein the primer sequence comprises a sequence configured for random priming.

6. The system of claim 1 , wherein the at least 1,000,000 oligonucleotide barcodes are attached to the bead via a chemical cross-linker.

7. The system of claim 6 , wherein the chemical cross-linker is a disulfide bond.

8. The system of claim 6 , wherein the at least 1,000,000 oligonucleotide barcodes are attached to the bead via a labile moiety.

9. The system of claim 1 , wherein the bead is degradable upon application of a stimulus.

10. The system of claim 9 , wherein the droplet further comprises the stimulus.

11. The system of claim 9 , wherein the stimulus is selected from the group consisting of a biological stimulus, a chemical stimulus, a thermal stimulus, an electrical stimulus, a magnetic stimulus, and a photo stimulus.

12. The system of claim 11 , wherein the stimulus is a chemical stimulus that is a reducing agent.

13. The system of claim 1 , wherein the droplet is an emulsion.

14. The system of claim 13 , wherein the emulsion is a water-in-oil emulsion.

15. The system of claim 1 , wherein the system further comprises the aqueous mixture and the immiscible continuous phase.

16. The system of claim 1 , further comprising the sample.

17. The system of claim 16 , wherein the sample comprises a cell comprising the nucleic acid analyte.

18. The system of claim 17 , wherein the sample comprises a plurality of cells including the cell.

19. The system of claim 1 , wherein the system further comprises the droplet.

20. The system of claim 19 , wherein the droplet comprises cell lysis reagents.

21. The system of claim 19 , wherein the droplet comprises reagents sufficient for conducting an amplification reaction.

22. The system of claim 19 , wherein the droplet comprises a polymerase.

23. The system of claim 1 , wherein one or more of the at least 1,000,000 oligonucleotide barcodes comprises a uracil.

Assignees

• 10X Genomics Inc

Inventors

• Hindson Benjamin
• Saxonov Serge
• Schnall-Levin Michael

Classifications

• C12Q2563/159

  Microreactors, e.g. emulsion PCR or sequencing, droplet PCR, microcapsules, i.e. non-liquid containers with a range of different permeability's for different reaction components · CPC title

• C12Q2535/122

  Massive parallel sequencing · CPC title

• B01L3/502715

  characterised by interfacing components, e.g. fluidic, electrical, optical or mechanical interfaces · CPC title

• B01J19/0046

  Sequential or parallel reactions, e.g. for the synthesis of polypeptides or polynucleotides; Apparatus and devices for combinatorial chemistry or for making molecular arrays (synthesis methods per se C40B50/00) · CPC title

• C12N15/1065

  Preparation or screening of tagged libraries, e.g. tagged microorganisms by STM-mutagenesis, tagged polynucleotides, gene tags · CPC title