Granted Patent B2

US 12,545,949 · App. 17/312,714 · Granted Feb 10, 2026

Inventors: Zachary Bent (Pleasanton, CA); Eswar Prasad Ramachandran Iyer (Sunnyvale, CA); Tarjei Sigurd Mikkelsen (Pleasanton, CA)

Assignee: 10x Genomics, Inc.

C12Q1/6844B01L3/50853B01L3/545B01L9/523C12N15/1065C12Q1/6837C12Q1/6841C12Q1/6874C12Q1/6876C12Q1/6881G01N21/6458G01N33/4833G01N35/00732G02B21/16G02B21/26G02B21/34G02B21/365B01L2300/0829C12Q1/6855C12Q1/6869C12Q2525/161C12Q2525/179C12Q2527/156C12Q2543/101C12Q2563/149C12Q2565/537C12Q2565/60C12Q2600/158G01N2021/6439G01N2035/00752

Abstract

Claims (30)

1 . A method for identifying a location of an analyte in a biological sample, the method comprising:

providing an array comprising a plurality of features on a substrate,

wherein a feature of the plurality of features comprises:

an oligonucleotide comprising a first spatial barcode; and

a capture probe comprising a second spatial barcode and a capture domain;

hybridizing a priming oligonucleotide to the oligonucleotide, wherein the priming oligonucleotide comprises a sequence that is substantially complementary to a portion of the oligonucleotide;

extending the priming oligonucleotide using the oligonucleotide as a template, thereby generating an extended priming oligonucleotide;

determining all or a portion of the sequence of the extended priming oligonucleotide and a location of the extended priming oligonucleotide;

determining a sequence and a location of the first spatial barcode, and a location of the second spatial barcode based on the location of the first spatial barcode;

associating the location of the extended priming oligonucleotide with the location of the first spatial barcode;

capturing the analyte from the biological sample with the capture domain; and

identifying the location of the analyte in the biological sample based on the location of the second spatial barcode.

2 . The method of claim 1 , wherein the oligonucleotide further comprises a cleavage domain.

3 . The method of claim 2 , wherein the cleavage domain comprises at least one of a chemical reagent-cleavable linker, a heat-cleavable linker, an ultrasound-cleavable linker, a photo-cleavable linker, and an enzyme-cleavable linker.

4 . The method of claim 1 , further comprising amplifying at least a portion of the oligonucleotide prior to determining the sequence of the first spatial barcode of the oligonucleotide.

5 . The method of claim 1 , wherein determining the sequence of the first spatial barcode comprises sequencing-by-synthesis.

6 . The method of claim 1 , wherein determining the sequence of the first spatial barcode comprises sequence-by-ligation.

7 . The method of claim 1 , wherein the capture probe further comprises a cleavage domain.

8 . The method of claim 1 , wherein the feature comprises a plurality of oligonucleotides and a plurality of capture probes, and wherein a ratio of oligonucleotides to capture probes on the feature is between 1:10 and 1:100,000.

9 . The method of claim 1 , wherein the first spatial barcode and the second spatial barcode are identical.

10 . The method of claim 1 , wherein the first spatial barcode and the second spatial barcode are different.

11 . The method of claim 1 , wherein the capture domain is complementary to a nucleic acid sequence present in or associated with the analyte.

12 . The method of claim 11 , wherein capturing the analyte from the biological sample with the capture domain comprises contacting the array with the biological sample and hybridizing the analyte to the capture domain.

13 . The method of claim 11 , wherein capturing the analyte from the biological sample with the capture domain comprises releasing the capture probe from the feature and contacting the released capture probe with the biological sample.

14 . The method of claim 1 , wherein the analyte comprises DNA or RNA.

15 . The method of claim 4 , wherein the amplifying comprises isothermal amplification.

16 . The method of claim 1 , wherein the capture probe further comprises a unique molecular identifier and one or more functional domains.

17 . The method of claim 1 , wherein the capture domain comprises a poly(T) sequence.

18 . The method of claim 1 , wherein the biological sample is a tissue section.

19 . The method of claim 18 , wherein the tissue section is a fresh-frozen tissue section or a fixed tissue section.

Assignments (1)

Continuity (38)

References Cited (400)

Cited By (2)

US 12,637,706 US 12,692,492