CREATION OF LIBRARIES OF DROPLETS AND RELATED SPECIES

• US 20150336071A1
• Filed: 07/29/2015
• Published: 11/26/2015
• Est. Priority Date: 09/19/2008
• Status: Active Grant

• Current Assignee: President and Fellows of Harvard College (Harvard Corporation)
• Original Assignee: President and Fellows of Harvard College (Harvard Corporation)
• Inventors: Weitz, David A., Agresti, Jeremy
• Granted Patent: US 11,401,550 B2
• Time in Patent Office: Days
• US Class Current: 1/1
• CPC Class Codes: B01J 13/04 by physical processes, e.g. drying, spraying
• CPC Class Codes: B01J 19/0046 Sequential or parallel reactions, e.g. for the synthesis of polypeptides or polynucleotides; Apparatus and devices for combinatorial chemistry or for making molecular arrays synthesis methods per se C40B50/00
• CPC Class Codes: B01J 2219/00599 Solution-phase processes
• CPC Class Codes: B01J 2219/00709 Type of synthesis
• CPC Class Codes: B01J 2219/00711 Light-directed synthesis
• CPC Class Codes: B01J 2219/00716 Heat activated synthesis
• CPC Class Codes: B01J 2219/00722 Nucleotides
• CPC Class Codes: C12Q 1/6874 involving nucleic acid arrays, e.g. sequencing by hybridisation
• CPC Class Codes: C12Q 1/6876 Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes

Abstract

The present invention is generally related to systems and methods for producing a plurality of droplets. The droplets may contain varying species, e.g., for use as a library. In some cases, the fluidic droplets may be rigidified to form rigidified droplets (e.g., gel droplets). In certain embodiments, the droplets may undergo a phase change (e.g., from rigidified droplets to fluidized droplets), as discussed more herein. In some cases, a species may be added internally to a droplet by exposing the droplet to a fluid comprising a plurality of species.

Claims

1-45. -45. (canceled)

46. A method, comprising:

providing a plurality of groups of rigidified droplets, each of the groups of rigidified droplets comprising a first fluid and having substantially the same composition as the other groups of rigidified droplets but containing a distinguishable species with respect to the other groups of rigidified droplets;

forming a suspension comprising at least one rigidified droplet from each of the groups of rigidified droplets;

exposing at least some of the rigidified droplets of the suspension to a second fluid, wherein the first fluid and the second fluid are substantially immiscible; and

fluidizing at least some of the rigidified droplets to form a plurality of fluidized droplets, wherein the plurality of fluidized droplets are substantially immiscible in the second fluid.

47. The method of claim 46, wherein the rigidified droplets are formed using microfluidic techniques.

48. The method of claim 46, wherein the first fluid is hydrophilic and the second fluid is hydrophobic.

49. The method of claim 46, wherein the first fluid is hydrophobic and the second fluid is hydrophilic.

50. The method of claim 46, wherein the plurality of groups of rigidified droplets comprises substantially of the same composition.

51. The method of claim 46, wherein the plurality of rigidified droplets comprise a polymer.

52. The method of claim 51, wherein the polymer comprises a plurality of crosslinks.

53. The method of claim 52, wherein the rigidified droplets are fluidized by causing the polymer crosslinks to cleave.

54. The method of claim 46, wherein the plurality of rigidified droplets comprise a gel.

55. The method of claim 54, wherein the wherein the rigidified droplets are caused to fluidize by causing the gel to reform a gel precursor.

56. The method of claim 46, wherein the rigidified droplets are fluidized by raising or lowering the temperature of the droplet.

57. The method of claim 56, wherein the temperature of the rigidified droplets is raised or lowered by about 10°

C.

58. The method of claim 56, wherein the temperature of the rigidified droplets is raised or lowered by about 20°

C.

59. The method of claim 46, wherein the rigidified droplets are fluidized by a changing the pH of the droplet.

60. The method of claim 46, further comprising the step of fusing at least some of the plurality of fluidized droplets with a plurality of microfluidic droplets to form a plurality of fused droplets.

61. The method of claim 60, wherein the plurality of microfluidic droplets comprise a target analyte molecule.

62. The method of claim 61, wherein the target analyte molecule is a nucleic acid.

63. The method of claim 61, further comprising the step of determining the target analyte molecule.

64. The method of claim 62, further comprising the step of determining the sequence of the nucleic acid.

65. The method of claim 46, wherein at least about 5 groups of rigidified droplets are provided.

66. The method of claim 46, wherein at least about 10 groups of rigidified droplets are provided.

67. The method of claim 46, wherein at least about 20 groups of rigidified droplets are provided.

68. The method of claim 46, wherein at least about 64 groups of rigidified droplets are provided.

69. The method of claim 46, wherein at least about 100 groups of rigidified droplets are provided.

70. The method of claim 46, wherein at least about 4096 groups of rigidified droplets are provided.

71. The method of claim 46, wherein at least about 10,000 groups of rigidified droplets are provided.

72. The method of claim 46, wherein the distinguishable species are nucleic acid probes.

73-155. -155. (canceled)