Systems and methods for sample analysis

Patent metadata

Field	Value
Publication number	US-12169198-B2
Application number	US-202318235616-A
Country	US
Kind code	B2
Filing date	Aug 18, 2023
Priority date	Jan 8, 2019
Publication date	Dec 17, 2024
Grant date	Dec 17, 2024

Abstract

The present disclosure provides systems and methods for processing or analyzing a sample. Methods may include generating supports (e.g., beads) comprising barcode molecules coupled thereto. A support comprising barcode molecules may be useful for analyzing or processing one or more analytes such as nucleic acid molecules, proteins, and/or perturbation agents.

First claim

What is claimed is:

1. A mixture, comprising:
a nucleic acid comprising: (i) a first moiety, (ii) a nucleic acid sequence, and (ii) a poly-thymine (poly(T)) sequence, a poly-guanine (poly(G)) sequence, or an N-mer sequence;
a nucleic acid barcode comprising a first strand and a second strand, wherein said first strand comprises a second moiety and a barcode sequence, wherein said second strand comprises (1) a first sequence complementary to said barcode sequence and (2) an overhang comprising a second sequence complementary to said nucleic acid sequence; and
reagents configured to non-enzymatically ligate said first moiety to said second moiety.

2. The mixture of claim 1 , further comprising a nucleic acid analyte, wherein said nucleic acid analyte is selected from the group consisting of a messenger ribonucleic acid (mRNA) molecule, a deoxyribonucleic acid (DNA) molecule, and an oligonucleotide attached to a labeling agent.

3. The mixture of claim 2 , wherein said nucleic acid analyte is said oligonucleotide attached to said labeling agent, and wherein said labeling agent is selected from the group consisting of a protein, antibody, and a lipid molecule.

4. The mixture of claim 2 , wherein said poly(T) sequence, said poly(G) sequence, or said N-mer sequence is part of a single stranded molecule configured to hybridize to a single-stranded end of a nucleic acid molecule of said nucleic acid analyte.

5. The mixture of claim 1 , wherein said first moiety and said second moiety are configured to generate a chemical bond selected from the group consisting of a triazole bond, a phosphorothioate bond, and an amide bond, when ligated together.

6. The mixture of claim 1 , further comprising a support, wherein said nucleic acid barcode is coupled to said support.

7. The mixture of claim 6 , wherein said support comprises a plurality of nucleic acid barcodes comprising said nucleic acid barcode.

8. The mixture of claim 6 , wherein said nucleic acid barcode is releasably coupled to said support.

9. The mixture of claim 8 , wherein said nucleic acid barcode is releasably coupled to said support by a labile bond, and wherein said labile bond is selected from the group consisting of a thermally labile bond, a chemically labile bond, and a photolabile bond.

10. The mixture of claim 6 , wherein said support is a bead.

11. The mixture of claim 10 , wherein said bead is a gel bead.

12. The mixture of claim 11 , wherein said gel bead is configured to degrade upon application of a stimulus.

13. The mixture of claim 12 , wherein said stimulus is selected from the group consisting of a thermal stimulus, a photo stimulus, a mechanical stimulus, a radiation stimulus, and a biological stimulus.

14. The mixture of claim 12 , further comprising a chemical stimulus.

15. The mixture of claim 1 , wherein said nucleic acid comprises said N-mer sequence located at a 3′ end of said nucleic acid.

16. The mixture of claim 1 , wherein said nucleic acid comprises said poly(G) sequence located at a 3′ end of said nucleic acid.

17. The mixture of claim 1 , wherein said nucleic acid is configured to be non-enzymatically ligated to a 5′ region of said first strand of said nucleic acid barcode.

18. The mixture of claim 1 , wherein said nucleic acid is configured to be non-enzymatically ligated to a 3′ region of said first strand of said nucleic acid barcode.

19. The mixture of claim 1 , wherein said nucleic acid comprises said poly(T) sequence located at a 3′ end of said nucleic acid.

20. The mixture of claim 1 , wherein said nucleic acid barcode further comprises one or more additional sequences selected from the group consisting of a sequence for permitting said nucleic acid barcode or a derivative thereof to couple to a flow cell of a sequencer, an adapter sequence, a unique molecular identifier sequence, a primer sequence, and a primer binding sequence.

21. The mixture of claim 1 , wherein said first moiety comprises an azide group and said second moiety comprises an alkyne group.

22. The mixture of claim 1 , wherein said first moiety comprises an alkyne group and said second moiety comprises an azide group.

Assignees

• 10X Genomics Inc

Inventors

• Price Andrew D
• Hindson Christopher

Classifications

• G01N33/58

  involving labelled substances (G01N33/53 takes precedence) · CPC title

• G01N33/56977

  HLA or MHC typing · CPC title

• G01N33/548

  Carbohydrates, e.g. dextran · CPC title

• G01N33/54366

  Apparatus specially adapted for solid-phase testing · CPC title

• G01N33/54306

  Solid-phase reaction mechanisms · CPC title