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US 2021/0230583 A1Published application
Barcoded wells for spatial mapping of single cells through sequencing
AssigneeBecton, Dickinson and Company
InventorGretchen Lam
Life of this patent
Estimating the term…
Abstract
Disclosed herein include systems, methods, compositions, and kits for methods of assigning sequencing data to partitions. There are provided, in some embodiments, methods of associating sequencing data and phenotypic data of single cells. There are provided, in some embodiments, methods of reducing noise in sequencing data. Disclosed herein include partition indexing oligonucleotides comprising a partition indexing sequence. The partition indexing oligonucleotides can be associated with partitions. Partition indexing oligonucleotides situated within the same partition can comprise the same partition indexing sequence. Partition indexing oligonucleotides situated within different partitions comprise different partition indexing sequences.
Claims20
A method of assigning sequencing data to partitions, comprising:
- providing a plurality of partitions each comprising a plurality of partition indexing oligonucleotides, wherein the partition indexing oligonucleotides each comprises a predetermined partition indexing sequence, wherein partition indexing oligonucleotides situated within the same partition comprise the same partition indexing sequence, and wherein partition indexing oligonucleotides situated within different partitions comprise different partition indexing sequences;
- partitioning a plurality of solid supports and a plurality of single cells to the plurality of partitions, wherein the single cells each comprises copies of a nucleic acid target, wherein the plurality of solid supports each comprises a plurality of oligonucleotide barcodes each comprising a cell label sequence, wherein oligonucleotide barcodes associated with the same solid support comprise the same cell label sequence, and wherein oligonucleotide barcodes associated with different solid supports comprise different cell label sequences;
- barcoding the partition indexing oligonucleotides using the plurality of oligonucleotide barcodes to generate a plurality of barcoded partition indexing oligonucleotides;
- barcoding the copies of a nucleic acid target from at least one of the plurality of single cells using the plurality of oligonucleotide barcodes to generate a plurality of barcoded nucleic acid targets;
- obtaining sequencing data comprising a plurality of sequencing reads of the barcoded partition indexing oligonucleotides, or products thereof, and the plurality of barcoded nucleic acid targets, or products thereof;
- identifying the partition indexing sequence associated with each cell label sequence in the sequencing data;
- and
- assigning each of the plurality of sequencing reads to a partition of the plurality of partitions based on the partitioning indexing sequence associated with each cell label sequence in the sequencing data.
5 more elements
A method of associating sequencing data and phenotypic data of single cells, comprising:
- obtaining phenotypic data of a plurality of single cells, wherein the single cells each comprises copies of a nucleic acid target;
- providing a plurality of partitions each comprising a plurality of partition indexing oligonucleotides, wherein the partition indexing oligonucleotides each comprises a predetermined partition indexing sequence, wherein partition indexing oligonucleotides situated within the same partition comprise the same partition indexing sequence, and wherein partition indexing oligonucleotides situated within different partitions comprise different partition indexing sequences;
- partitioning each of the plurality of single cells to an identified partition of the plurality of partitions;
- barcoding the copies of a nucleic acid target from at least one of the plurality of single cells using a plurality of oligonucleotide barcodes associated with a solid support to generate a plurality of barcoded nucleic acid targets;
- barcoding the partition indexing oligonucleotides using a plurality of oligonucleotide barcodes associated with a solid support to generate a plurality of barcoded partition indexing oligonucleotides;
- obtaining sequencing data of the plurality of barcoded partition indexing oligonucleotides, or products thereof, and the plurality of barcoded nucleic acid targets, or products thereof;
- and
- associating the sequencing data and the phenotypic data of at least one cell of the plurality of single cells based on the partition indexing sequence of at least one barcoded partition indexing oligonucleotide, or product thereof, of the plurality of barcoded partition indexing oligonucleotides, or products thereof, in the sequencing data.
5 more elements
A composition comprising:
- a micro-well array,
- wherein the micro-well array comprises at least 100 micro-wells, wherein each micro-well has a volume ranging from about 1,000 μm3 to about 786,000 μm3, wherein each micro-well comprises a plurality of partition indexing oligonucleotides, wherein the partition indexing oligonucleotides each comprises a predetermined partition indexing sequence, wherein partition indexing oligonucleotides situated within the same micro-well comprise the same partition indexing sequence, and wherein partition indexing oligonucleotides situated within different micro-wells comprise different partition indexing sequences.
Dependencies are inferred from the claim text.