Methods and systems for processing polynucleotides
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.
1. A system for nucleic acid processing, comprising:
a microwell array comprising 1,000 microwells configured to partition a plurality of polynucleotide molecules, a plurality of beads, and a plurality of barcode molecules in the 1,000 microwells of the microwell array,
wherein the plurality of barcode molecules comprises a plurality of barcode sequences that are different across the 1,000 microwells, and
wherein a microwell of the microwell array comprises: (i) a polynucleotide molecule from the plurality of polynucleotide molecules; and (ii) a barcode molecule from the plurality of barcode molecules, wherein the barcode molecule is attached to a bead from the plurality of beads and comprises a barcode sequence from the plurality of barcode sequences.
2. The system of claim 1 , further comprising a sequencer that generates sequences of the plurality of polynucleotide molecules or derivatives thereof.
3. The system of claim 1 , wherein the plurality of beads is a plurality of solid particles.
4. The system of claim 1 , wherein the plurality of beads is a plurality of gel beads.
5. The system of claim 1 , wherein the microwell comprises a cell comprising the polynucleotide molecule from the plurality of polynucleotide molecules.
6. The system of claim 1 , wherein the microwell comprises a reducing agent.
7. The system of claim 1 , wherein the barcode molecule comprises a sequence complementary to a sequence of the polynucleotide molecule.
8. The system of claim 1 , wherein the plurality of polynucleotide molecules comprises polynucleotide molecules with overlapping sequences.
9. The system of claim 1 , wherein the microwell comprises an antibody.
10. The system of claim 1 , wherein the microwell comprises multiple barcode molecules from the plurality of barcode molecules, wherein the multiple barcode molecules comprise the barcode molecule.
11. The system of claim 10 , wherein each of the multiple barcode molecules comprises the barcode sequence.
12. The system of claim 1 , wherein the 1,000 microwells are a subset of microwells of the microwell array.
13. The system of claim 12 , wherein at least one microwell of the microwell array does not include a barcode molecule from the plurality of barcode molecules or a polynucleotide molecule from the plurality of polynucleotide molecules.
14. The system of claim 1 , wherein the microwell of the microwell array is configured to accommodate a volume of less than about 500 nanoliters (nL).
15. The system of claim 1 , wherein the microwell array comprises at least 5,000 microwells.
16. A system for nucleic acid processing, comprising:
a microwell array comprising a plurality of microwells configured to partition a plurality of beads and a plurality of cells;
wherein the plurality of microwells comprises 1,000 microwells;
wherein a microwell of the plurality of microwells is configured to accommodate a volume of less than about 5 nanoliters (nL); and
wherein the system is configured to partition the plurality of cells, the plurality of beads, and a plurality of barcode molecules into the plurality of microwells such that the plurality of barcode molecules comprises a plurality of barcode sequences that are different across microwells of the microwell array, and wherein the microwell is configured to comprise: (a) a cell of the plurality of cells; and (b) a bead of the plurality of beads.
17. The system of claim 16 , wherein the plurality of barcode molecules is attached to the plurality of beads.
18. The system of claim 17 , wherein the bead of (b) is attached to a barcode molecule of the plurality of barcode molecules and comprises a barcode sequence that is different from barcode sequences of other barcode molecules attached to other beads in the microwells.
19. The system of claim 16 , further comprising a sequencer that generates sequences of the plurality of barcode molecules or derivatives thereof.
20. The system of claim 16 , wherein the plurality of beads is a plurality of solid particles.
21. The system of claim 16 , wherein the plurality of beads is a plurality of gel beads.
22. The system of claim 16 , wherein the microwell further comprises a reducing agent.
23. The system of claim 16 , wherein the microwell of the microwell array comprises an antibody.
24. The system of claim 16 , wherein the microwell array comprises 10,000 microwells and wherein the microwell is configured to comprise: (a) the cell of the plurality of cells; and (b) the bead of the plurality of beads.
25. The assay of claim 16 , wherein at least one microwell of the plurality of microwells does not include a barcode molecule from the plurality of barcode molecules or a cell from the plurality of cells.
26. A microwell array, comprising:
a plurality of microwells comprising a plurality of cells and a plurality of beads;
wherein the plurality of beads comprises a plurality of barcode molecules;
wherein the plurality of microwells comprises 1,000 microwells;
wherein the plurality of barcode molecules comprise a plurality of barcode sequences that are different across the 1,000 microwells;
wherein a microwell of the plurality of microwells is configured to accommodate a volume of less than about 5 nanoliters; and
wherein the microwell comprises: (a) a cell of the plurality of cells; and (b) a bead of the plurality of beads, wherein the bead is attached to a barcode molecule of the plurality of barcode molecules.
27. The microwell array of claim 26 , wherein the barcode molecule attached to the bead of (b) comprises a barcode sequence that is different from barcode sequences of barcode molecules attached to other beads in microwells of the plurality of microwells.
28. The microwell array of claim 26 , wherein the plurality of beads is a plurality of solid particles.
29. The microwell array of claim 26 , wherein the plurality of beads is a plurality of gel beads.
30. The microwell array of claim 26 , wherein the microwell comprises a reducing agent.
31. The microwell array of claim 26 , wherein the microwell comprises an antibody.
32. The microwell array of claim 26 , wherein at least one microwell does not include a barcode molecule from the plurality of barcode molecules or a cell from the plurality of cells.
33. The microwell array of claim 26 , wherein the plurality of microwells comprises 10,000 microwells and wherein the microwell comprises: (a) a cell of the plurality of cells; and (b) a bead of the plurality of beads.
34. The system of claim 1 , wherein the polynucleotide molecule does not include a barcode sequence, and wherein the polynucleotide molecule is hybridized to the barcode molecule.
35. The system of claim 1 , wherein the bead in the microwell is not attached to an amplification product of the polynucleotide molecule.
36. The system of claim 18 , wherein the cell comprises a plurality of polynucleotide molecules, and wherein the barcode sequence comprises a sequence that is complementary to a sequence of a polynucleotide of the plurality of polynucleotide molecules.
37. The system of claim 14 , wherein the microwell is configured to accommodate a volume of less than about 200 nL.
38. The system of claim 37 , wherein the microwell is configured to accommodate a volume of less than about 100 nL.
39. The system of claim 38 , wherein the microwell is configured to accommodate a volume of less than about 50 nL.
40. The system of claim 39 , wherein the microwell is configured to accommodate a volume of less than about 25 nL.
41. The system of claim 40 , wherein the microwell is configured to accommodate a volume of less than about 10 nL.
42. The system of claim 41 , wherein the microwell is configured to accommodate a volume of less than about 5 nL.
43. The system of claim 15 , wherein the microwell array comprises 2,500 microwells.
44. The system of claim 43 , wherein the microwell array comprises 5,000 microwells.
45. The system of claim 44 , wherein the microwell array comprises 7,500 microwells.
46. The system of claim 45 , wherein the microwell array comprises 10,000 microwells.
47. The system of claim 16 , wherein the microwell comprises the cell of the plurality of cells and the bead of the plurality of beads.