METHODS AND SYSTEMS FOR PROCESSING POLYNUCLEOTIDES

• US 20180282804A1
• Filed: 06/05/2018
• Published: 10/04/2018
• Est. Priority Date: 12/14/2012
• Status: Abandoned Application

• Current Assignee: 10X Genomics Incorporated
• Original Assignee: 10X Genomics Incorporated
• Inventors: Hindson, Benjamin, Saxonov, Serge, Ness, Kevin, Hardenbol, Paul, Schnall-Levin, Michael, Jarosz, Mirna
• Application Number:
  US16/000,803
• Publication Number:
  US 20180282804A1
• Time in Patent Office: Days
• CPC Class Codes: C12Q 1/6806 Preparing nucleic acids for analysis, e.g. for polymerase chain reaction [PCR] assay C12Q1/6804 takes precedence
• CPC Class Codes: C12Q 1/6855 Ligating adaptors
• CPC Class Codes: C12Q 1/6869 Methods for sequencing
• CPC Class Codes: C12Q 1/6874 involving nucleic acid arrays, e.g. sequencing by hybridisation
• CPC Class Codes: C12Q 2525/161 incorporating target specific and non-target specific sites
• CPC Class Codes: C12Q 2535/122 Massive parallel sequencing
• CPC Class Codes: C12Q 2563/159 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 Class Codes: C12Q 2563/179 the label being a nucleic acid
• CPC Class Codes: C12Q 2565/514 characterised by the use of the arrayed oligonucleotides as identifier tags, e.g. universal addressable array, anti-tag or tag complement array

Abstract

The present disclosure provides compositions, methods, systems, and devices for polynucleotide processing. Such polynucleotide processing may be useful for a variety of applications, including polynucleotide sequencing.

Claims

1. A method for nucleic acid preparation or analysis, comprising:

(a) partitioning a plurality of barcode molecules and a plurality of nucleic acid molecules derived from a cell-free biological sample in a plurality of partitions, wherein said plurality of barcode molecules comprise a plurality of barcode sequences;

(b) in said plurality of partitions, using said plurality of nucleic acid molecules and said plurality of barcode molecules to generate a plurality of barcoded nucleic acid molecules, which plurality of barcoded nucleic acid molecules comprises said plurality of barcode sequences;

(c) recovering or releasing said plurality of barcoded nucleic acid molecules from said plurality of partitions.

2. The method of claim 1, wherein a barcode sequence associated with a given partition of said plurality of partitions is different from a barcode sequence associated with other partitions of said plurality of partitions.

3. The method of claim 1, wherein said plurality of partitions is a plurality of droplets.

4. The method of claim 1, wherein said plurality of partitions is a plurality of wells.

5. The method of claim 1, wherein said plurality of barcode molecules are coupled to beads.

6. The method of claim 5, wherein said beads are solid particles.

7. The method of claim 5, wherein said beads are gel beads.

8. The method of claim 5, further comprising releasing said plurality of barcode molecules from said beads in said plurality of partitions.

9. The method of claim 5, wherein said plurality of barcode molecules is covalently attached to said beads through linkers.

10. The method of claim 1, wherein said plurality of nucleic acid molecules are ribonucleic acid molecules.

11. The method of claim 1, wherein said plurality of nucleic acid molecules are deoxyribonucleic acid molecules.

12. The method of claim 1, wherein said cell-free biological sample is a bodily fluid sample.

13. The method of claim 12, wherein said bodily fluid sample is blood.

14. The method of claim 12, wherein said bodily fluid sample is serum.

15. The method of claim 12, wherein said bodily fluid sample is saliva.

16. The method of claim 12, wherein said bodily fluid sample is urine.

17. The method of claim 1, wherein in (c), said plurality of barcoded nucleic acid molecules is generated by nucleic acid amplification using said plurality of nucleic acid molecules and said plurality of barcode molecules.

18. The method of claim 1, wherein a partition of said plurality of partitions comprises barcode molecules from said plurality of barcode molecules, and wherein said barcode molecules comprise a common barcode sequence.

19. The method of claim 1, further comprising, subsequent to (c), sequencing said plurality of barcoded nucleic acid molecules or derivatives thereof to generate a plurality of sequences, which plurality of sequences comprise sequences of said plurality of nucleic acid molecules derived from said cell-free biological sample and said plurality of barcode sequences.

20. The method of claim 19, further comprising using said plurality of sequences to differentiate between haplotypes in said plurality of nucleic acid molecules derived from said cell-free biological sample.