# US 10,767,223 B1 · Methods for analyzing nucleic acids from single cells

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- Number: US 10,767,223 B1
- Document ID: US10767223B1
- Status: Granted
- Kind: B1
- Application number: 15/930,333
- Filed: 2020-05-12
- Published: 2020-08-27
- Granted: 2020-09-08
- Earliest claimed: 2009-08-20
- Assignees: 10x Genomics, Inc.
- Applicants: 10X Genomics, Inc.
- Inventors: Sydney Brenner; Gi Mikawa; Robert Osborne; Andrew Slatter
- Web page: https://patentgrep.com/patent/US10767223B1

## CPC

- C12Q 1/6874: involving nucleic acid arrays, e.g. sequencing by hybridisation
- C12N 15/1065: {Preparation or screening of tagged libraries, e.g. tagged microorganisms by STM-mutagenesis, tagged polynucleotides, gene tags}
- C12Q 1/6806: Preparing nucleic acids for analysis, e.g. for polymerase chain reaction [PCR] assay (C12Q1/6804 takes precedence)
- C12Q 1/6855: Ligating adaptors
- C12Q 1/686: Polymerase chain reaction [PCR]
- C12Q: MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS (immunoassay G01N33/53); COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
- C12N: MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA (microbiological testing media C12Q1/00)

## Abstract

Aspects of the present invention include analyzing nucleic acids from single cells using methods that include using tagged polynucleotides containing multiplex identifier sequences.

## Claims

### Claim 1

A method for biological analysis, the method comprising:
(a) providing a plurality of polynucleotides derived from a tissue sample;
(b) generating a plurality of tagged polynucleotides from said plurality of polynucleotides and a plurality of oligonucleotide tags, wherein a tagged polynucleotide of the plurality of tagged polynucleotides comprises:
(i) a first tag sequence corresponding to a site within said tissue sample; and
(ii) a second tag sequence distinguishing said tagged polynucleotide from other tagged polynucleotides of the plurality of tagged polynucleotides;
(c) sequencing said plurality of tagged polynucleotides to determine sequences corresponding to the plurality of polynucleotides, including the first tag sequence and the second tag sequence; and
(d) using the sequences determined in step (c) to count polynucleotides for multiple different polynucleotides derived from said tissue sample.

### Claim 2 (depends on 1)

The method of claim 1, wherein said plurality of polynucleotides is provided in a well of a plurality of wells.

### Claim 3 (depends on 1)

The method of claim 1, wherein an oligonucleotide tag of said plurality of oligonucleotide tags further comprises a sequence that is configured to hybridize to said plurality of polynucleotides.

### Claim 4 (depends on 1)

The method of claim 1, wherein substantially every polynucleotide of said plurality of polynucleotides is associated with the same first tag sequence.

### Claim 5 (depends on 1)

The method of claim 1, wherein an oligonucleotide tag of said plurality of oligonucleotide tags comprises a sequencing adaptor.

### Claim 6 (depends on 5)

The method of claim 5, wherein step (c) comprises hybridizing said plurality of tagged polynucleotides or derivatives thereof to a solid support via said sequencing adaptor or derivative thereof.