Methods and systems for processing polynucleotides

Patent metadata

Field	Value
Publication number	US-2018094312-A1
Application number	US-201715831726-A
Country	US
Kind code	A1
Filing date	Dec 5, 2017
Priority date	Jun 26, 2014
Publication date	Apr 5, 2018
Grant date	—

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.

First claim

What is claimed is:

1 . A library of gel beads for barcoding nucleic acid molecules, comprising:
a plurality of gel beads that are degradable upon application of a stimulus, wherein said plurality of gel beads comprises a plurality of barcode sequences that are different across said plurality of gel beads, wherein a given gel bead of said plurality of gel beads comprises a plurality of nucleic acid molecules, wherein each of said plurality of nucleic acid molecules comprises a barcode sequence from said plurality of barcode sequences and a unique molecular sequence, wherein (i) said barcode sequence is constant across said plurality of nucleic acid molecules, and (ii) said unique molecular sequence varies across said plurality of nucleic acid molecules.

2 . The library of claim 1 , wherein said plurality of gel beads comprises at least 1,000 gel beads.

3 . The library of claim 2 , wherein said plurality of gel beads comprises at least 10,000 gel beads.

4 . The library of claim 3 , wherein said plurality of gel beads comprises at least 100,000 gel beads.

5 . The library of claim 1 , wherein said plurality of nucleic acid molecules comprises at least 1,000 nucleic acid molecules.

6 . The library of claim 5 , wherein said plurality of nucleic acid molecules comprises at least 10,000 nucleic acid molecules.

7 . The library of claim 6 , wherein said plurality of nucleic acid molecules comprises at least 100,000 nucleic acid molecules.

8 . The library of claim 7 , wherein said plurality of nucleic acid molecules comprises at least 1,000,000 nucleic acid molecules.

9 . The library of claim 1 , wherein each of said plurality of nucleic acid molecules comprises a functional sequence for coupling to a flow cell of a sequencer.

10 . The library of claim 1 , wherein each of said plurality of nucleic acid molecules comprises a priming sequence that is targeted to a nucleic acid molecule.

11 . The library of claim 10 , wherein said priming sequence is a poly-T sequence.

12 . The library of claim 11 , wherein each of said plurality of nucleic acid molecules further comprises an anchoring sequence to permit said poly-T sequence to hybridize at a sequence end of a target ribonucleic acid molecule.

13 . The library of claim 1 , wherein each of said plurality of nucleic acid molecules comprises a random priming sequence.

14 . The library of claim 1 , wherein said given gel bead is porous.

15 . The library of claim 1 , wherein said plurality of gel beads have substantially monodisperse cross-sectional dimensions.

16 . The library of claim 1 , wherein said plurality of nucleic acid molecules is coupled to said given gel bead.

17 . The library of claim 16 , wherein said plurality of nucleic acid molecules is releasably coupled to said given gel bead.

18 . A composition for barcoding a nucleic acid molecule, comprising:
a gel bead that is degradable upon application of a stimulus, wherein said gel bead comprises a plurality of nucleic acid molecules, wherein each of said plurality of nucleic acid molecules comprises a barcode sequence and a unique molecular sequence, wherein (i) said barcode sequence is constant across said plurality of nucleic acid molecules, and (ii) said unique molecular sequence varies across said plurality of nucleic acid molecules.

19 . The composition of claim 18 , wherein said plurality of nucleic acid molecules comprises at least 1,000 nucleic acid molecules.

20 . The composition of claim 19 , wherein said plurality of nucleic acid molecules comprises at least 10,000 nucleic acid molecules.

21 . The composition of claim 20 , wherein said plurality of nucleic acid molecules comprises at least 100,000 nucleic acid molecules.

22 . The composition of claim 21 , wherein said plurality of nucleic acid molecules comprises at least 1,000,000 nucleic acid molecules.

23 . The composition of claim 18 , wherein each of said plurality of nucleic acid molecules comprises a functional sequence for coupling to a flow cell of a sequencer.

24 . The composition of claim 18 , wherein each of said plurality of nucleic acid molecules comprises a priming sequence for a target nucleic acid molecule.

25 . The composition of claim 24 , wherein said priming sequence is a poly-T sequence.

26 . The composition of claim 25 , wherein each of said plurality of nucleic acid molecules further comprises an anchoring sequence to permit said poly-T sequence to hybridize at a sequence end of a target ribonucleic acid molecule.

27 . The composition of claim 18 , wherein each of said plurality of nucleic acid molecules comprises a random priming sequence.

28 . The composition of claim 18 , wherein said gel bead is porous.

29 . The composition of claim 18 , wherein said plurality of nucleic acid molecules is coupled to said gel bead.

30 . The composition of claim 29 , wherein said plurality of nucleic acid molecules is releasably coupled to said gel bead.

Assignees

• 10X Genomics Inc

Inventors

• Hindson Benjamin
• Hindson Christopher
• Schnall-Levin Michael
• Ness Kevin
• Jarosz Mirna
• Saxonov Serge
• Hardenbol Paul
• Bharadwaj Rajiv
• Zheng Xinying
• Belgrader Phillip

Classifications

• C12Q2537/143

  Multiplexing, i.e. use of multiple primers or probes in a single reaction, usually for simultaneously analyse of multiple analysis · CPC title

• C12Q1/6804

  Nucleic acid analysis using immunogens (immunoassay G01N33/53) · CPC title

• C12Q2563/179

  the label being a nucleic acid · CPC title

• C12Q2565/629

  being a microfluidic device · CPC title

• C12Q1/6874 Primary

  involving nucleic acid arrays, e.g. sequencing by hybridisation · CPC title