Chemical synthesis using solvent microdroplets

• US 6,419,883 B1
• Filed: 01/16/1998
• Issued: 07/16/2002
• Est. Priority Date: 01/16/1998
• Status: Expired due to Term

• Current Assignee: Washington University In St Louis
• Original Assignee: University of Washington
• Inventors: Blanchard, Alan P.
• Primary Examiner(s): Wilson, James O.
• Application Number:
  US09/008,120
• Time in Patent Office: 1,642 Days
• Field of Search: 422/102.2, 422/111, 422/149.11, 422/129.1, 422/131, 422/134, 422/136
• US Class Current: 422/134
• 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 2219/00378 Piezoelectric or ink jet dispensers
• CPC Class Codes: B01J 2219/00475 Sheets
• CPC Class Codes: B01J 2219/00488 by rotation of the reaction vessels
• CPC Class Codes: B01J 2219/00527 Sheets
• CPC Class Codes: B01J 2219/00536 in the shape of disks
• CPC Class Codes: B01J 2219/00585 Parallel processes
• CPC Class Codes: B01J 2219/0059 Sequential processes
• CPC Class Codes: B01J 2219/00596 Solid-phase processes
• CPC Class Codes: B01J 2219/00605 the compounds being directly bound or immobilised to solid supports
• CPC Class Codes: B01J 2219/00612 the surface being inorganic
• CPC Class Codes: B01J 2219/00619 using hydrophilic or hydrophobic regions
• CPC Class Codes: B01J 2219/00626 Covalent
• CPC Class Codes: B01J 2219/00659 Two-dimensional arrays
• CPC Class Codes: B01J 2219/00675 In-situ synthesis on the substrate
• CPC Class Codes: B01J 2219/00689 using computers
• CPC Class Codes: B01J 2219/00691 using robots
• CPC Class Codes: B01J 2219/00695 Synthesis control routines, e.g. using computer programs
• CPC Class Codes: B01J 2219/00704 integrated with the reactor apparatus
• CPC Class Codes: B01J 2219/00722 Nucleotides
• CPC Class Codes: B41J 2/01 : Ink jet
• CPC Class Codes: B82Y 30/00 : Nanotechnology for materials or surface science, e.g. nanocomposites
• CPC Class Codes: C07H 19/00 : Compounds containing a hetero ring sharing one ring hetero atom with a saccharide radical; Nucleosides; Mononucleotides; Anhydro-derivatives thereof
• CPC Class Codes: C07H 21/00 : Compounds containing two or more mononucleotide units having separate phosphate or polyphosphate groups linked by saccharide radicals of nucleoside groups, e.g. nucleic acids
• CPC Class Codes: C07K 1/02 : in solution C07K1/003, C07K1/006 take precedence
• CPC Class Codes: C07K 1/04 : on carriers C07K1/003, C07K1/006 take precedence
• CPC Class Codes: C40B 40/06 : Libraries containing nucleotides or polynucleotides, or derivatives thereof
• CPC Class Codes: C40B 50/14 : Solid phase synthesis, i.e. wherein one or more library building blocks are bound to a solid support during library creation; Particular methods of cleavage from the solid support

Abstract

The present invention relates to microdroplets of a solution comprising a solvent having a boiling point of 150° C. or above, a surface tension of 30 dynes/cm or above, and a viscosity of 0.015 g/(cm)(sec). Such microdroplets are useful for the synthesis of chemical species, particularly biopolymers such as oligonucleotides and peptides, as well as arrays of chemical species. Preferably, the solvent has the formula (I):

wherein

A=O or S;

X=O, S or N (C₁-C₄ alkyl)

Y=O, S, N(C₁-C₄ alkyl) or CH₂; and

R=C₁-C₂₀ straight or branched chain alkyl.

Claims

1. An automated system capable of processing one or more substrates, comprising:

an inkjet print head for spraying a microdroplet comprising a chemical species on a substrate;

a scanning transport for scanning the substrate adjacent to the print head to selectively deposit the microdroplet at specified sites;

a flow cell for treating the substrate on which the microdroplet is deposited by exposing the substrate to one or more selected fluids;

a treating transport for moving the substrate between the print head and the flow cell for treatment in the flow cell, wherein said treating transport and said scanning transport are different elements; and

an alignment unit for aligning the substrate correctly relative to the print head each time when the substrate is positioned adjacent to the print head for deposition.

2. The system of claim 1, wherein the inkjet printhead contains a solution comprising the chemical species dissolved in a solvent.

3. The system of claim 2, wherein the chemical species is a monomer unit of a biopolymer.

4. The system of claim 2, wherein the solution further comprises a catalyst.

5. The system of claim 2 wherein the solution comprises a solvent having a boiling point of 150°

C. or above, a surface tension of 30 dynes/cm or above, and a viscosity of 0.015 g/(cm)(sec) or above.

6. The system of claim 5, wherein the solvent has the formula (I):

wherein

A=O or S;

X=O, S or N (C₁-C₄ alkyl);

Y=O, S, N (C₁-C₄ alkyl) or CH₂; and

R=C₁-C₂₀ straight or branched chain alkyl.

7. The system of claim 6, wherein the solvent is selected from the group consisting of:

N-methyl-2-pyrrolidone;

2-pyrrolidone;

propylene carbonate;

γ

-valerolactone;

6-caprolactam;

ethylene carbonate;

γ

-butyrolactone;

δ

-valerolactone;

1,3-dimethyl-3,4,5,6-tetrahydro-2(1H)-pyrimidinone;

ethylene trithiocarbonate; and

1,3-dimethyl-2-imidazolidinone.

8. The system of claim 5, wherein said solvent is propylene carbonate.

9. The system of claim 3, which is for synthesizing an oligonucleotide, and wherein the monomer is a nucleoside or nucleoside derivative.

10. The system of claim 9, wherein the nucleoside is a deoxyribonucleoside or a ribonucleoside.

11. The system of claim 1, wherein the inkjet print head comprises an array of piezoelectric pumps.

12. The system of claim 11, further comprising an external reservoir connected to supply the chemical species to the print head.

13. The system of claim 12, wherein the external reservoir contains a solution comprising the chemical species dissolved in a solvent.

14. The system of claim 13, wherein the solvent has a boiling point of 150°

above, a surface tension of 30 dynes/cm or above, and a viscosity of 0.015 g/(cm)(sec) or above.

15. The system of claim 14, wherein the solvent has the formula (I):

wherein

A=O or S;

X=O, S or N (C₁-C₄ alkyl);

Y=O, S, N(C₁-C₄ alkyl) or CH₂; and

R=C₁-C₂₀ straight or branched chain alkyl.

16. The system of claim 15, wherein the solvent is selected from the group consisting of:

N-methyl-2-pyrrolidone;

2-pyrrolidone;

propylene carbonate;

γ

-valerolactone;

6-caprolactam;

ethylene carbonate;

γ

-butyrolactone;

δ

-valerolactone;

1 3-dimethyl-3,4,5,6-tetrahydro-2(1H)-pyrimidinone;

ethylene trithiocarbonate; and

1,3-dimethyl-2-imidazolidinone.

17. The system of claim 15, wherein said solvent is propylene carbonate.

83. The system of claim 1 or 18, wherein the substrate is porous.

89. The automated system of claim 1, further comprising a plurality of flow cells.

18. An automated system for synthesizing oligonucleotides on a substrate, said automated system capable of processing one or more substrates, comprising:

an inkjet print head for spraying a solution comprising a nucleoside or activated nucleoside on a substrate;

a scanning transport for scanning the substrate adjacent to the print head to selectively deposit the nucleoside at specified sites;

a flow cell for treating the substrate on which the monomer is deposited by exposing the substrate to one or more selected fluids;

a treating transport for moving the substrate between the print head and the flow cell for treatment in the flow cell, wherein said treating transport and said scanning transport are different elements; and

an alignment unit for aligning the substrate correctly relative to the print head each time when the substrate is positioned adjacent to the print head for deposition.

19. The system of claim 18, wherein the inkjet print head comprises an array of piezoelectric pumps.

20. The system of claim 18, further comprising:

an external reservoir connected to supply the nucleoside to the print head.

21. The system of claim 20, wherein the external reservoir contains a solution comprising said nucleoside or activated nucleoside dissolved in propylene carbonate.

22. The system of claim 18, further comprising a plurality of external reservoirs connected to the printer head, each external reservoir storing a nucleoside or activated nucleoside.

23. The system of claim 18, further comprising control logic configured to perform the following steps:

moving the substrate over the print head with the scanning transport;

firing the print head repeatedly to deposit the nucleoside or activated nucleoside monomer at the specified loci on the substrate; and

transferring the substrate to the flow cell with the treating transport.

24. The system of claim 18, wherein the scanning transport comprises:

a vacuum chuck for holding the substrate; and

a translational stage connected to move the vacuum chuck with respect to the print head.

25. The system of claim 24, wherein the vacuum chuck is rotatable for alignment with the print head.

26. The system of claim 25, wherein the vacuum chuck is engageable by a stationary element to rotate the vacuum chuck for alignment with the print head.

27. The system of claim 24, wherein the translational stage is driven by motorized means.

28. The system of claim 27, wherein the motorized means is a stepping motor.

29. The system of claim 18, wherein the flow cell has means for rinsing off unconnected monomers.

30. The system of claim 18, wherein the treating transport comprises:

a vacuum chuck for holding the substrate; and

a translational stage connected to move the second vacuum chuck and to move the substrate to and from the flow cell.

31. The system of claim 30, wherein the translational stage is driven by motorized means.

32. The system of claim 31, wherein the motorized means is a stepping motor.

33. The system of claim 18, wherein said alignment unit comprises a camera positioned adjacent to the substrate to positionally calibrate the substrate.

34. The system of claim 18, wherein said alignment unit comprises a marker that can be activated to establish one or more marks at particular loci on the substrate for positionally calibrating the substrate.

35. The system of claim 18, wherein said alignment unit comprises:

a marker that can be activated to establish one or more marks at particular loci on the substrate; and

a camera positioned adjacent to the substrate to locate said marks relative to the printer head.

36. The system of claim 35, further comprising a tip that can be activated to scratch marks at particular loci on the substrate for positionally calibrating the substrate.

37. The system of claim 18, wherein said alignment unit comprises:

control logic connected to control the movement of the scanning transport;

a marker that can be activated to establish one or more marks at particular loci on the substrate; and

a camera positioned adjacent to the substrate to locate said marks relative to the printer head.

38. The system of claim 37, wherein the control logic is configured to perform the following steps:

moving the substrate over the marker to establish one or more marks on the substrate;

subsequently locating the marks with the camera;

determining the position of the substrate with respect to the print head with reference to the marks; and

calibrating the scanning transport in response to the determined position of the substrate with respect to the printer head.

39. The system of claim 37, further comprising a stationary element that engages the substrate chuck to rotate the substrate chuck for alignment with the print head, wherein the control logic is configured to perform the following steps:

moving the substrate over the marker to establish one or more marks on the substrate;

subsequently locating the marks with the camera;

determining misalignment of the substrate relative to the print head with reference to the marks; and

moving the translational stage to (a) engage the substrate chuck with the stationary element, and (b) rotate the substrate chuck by an angular displacement that corrects for the misalignment.

40. The system of claim 18, further comprising a transfer station that supports the substrate for transfer between the treating transport and the scanning transport.

90. The automated system of claim 18, further comprising a plurality of flow cells.

41. A method of controlling a system for synthesizing a biopolymer on a substrate, said system capable of processing one or more substrates, using a computer having a memory for storing a control program and data, wherein the system comprises an inkjet print head for spraying a microdroplet on the substrate, a scanning transport for scanning the substrate adjacent to the print head to selectively deposit the microdroplet, an alignment unit for detecting misalignment of the substrate with respect to the print head at each deposition step, a flow cell for treating the substrate, and a treating transport for moving the substrate between the printer head and the flow cell, wherein said treating transport and said scanning transport are different elements, the method comprising the steps of:

(a) aligning the substrate relative to the print head by processing data from the alignment unit and by sending a signal to the scanning transport to move the substrate so as to correct misalignment of the substrate;

(b) selectively depositing a microdroplet on the substrate by sending a signal to the print head to spray the microdroplet and by sending a signal to the scanning transport to move the substrate adjacent to the print head so that the microdroplet can be deposited at specified loci on the substrate; and

(c) controlling treatment of the substrate by sending a signal to the treating transport to move the substrate to the flow cell and by sending a signal to the flow cell to control operation of the flow cell.

42. The method of claim 41, wherein the microdroplet comprises a monomer unit of a biopolymer.

43. The method of claim 42, wherein the microdroplet further comprises a catalyst.

44. The method of claim 41, wherein the microdroplet comprises a solvent.

45. The method of claim 44, wherein the solvent has a boiling point of 150°

C. or above, a surface tension of 30 dynes/cm or above, and a viscosity of 0.015 g/(cm)(sec) or above.

46. The method of claim 45, wherein the solvent has the formula (I):

wherein

A=O or S;

X=O, S or N (C₁-C₄ alkyl);

Y=O, S, N(C₁-C₄ alkyl) or CH₂; and

R=C₁-C₂₀ straight or branched chain alkyl.

47. The method of claim 46, wherein the solvent is selected from the group consisting of:

N-methyl-2-pyrrolidone;

2-pyrrolidone;

propylene carbonate;

γ

-valerolactone;

6-caprolactam;

ethylene carbonate;

γ

-butyrolactone;

δ

-valerolactone;

1,3-dimethyl-3,4,5,6-tetrahydro-2(1H)-pyrimidinone;

ethylene trithiocarbonate; and

1,3-dimethyl-2-imidazolidinone.

48. The method of claim 44, wherein said solvent is propylene carbonate.

49. The method of claim 41, further comprising the steps of repeating said steps (a)-(c).

50. The method of claim 42, further comprising the steps of repeating said steps (a)-(c) to form biopolymer array.

51. The method of claim 41, wherein the biopolymer is an oligonucleotide.

52. The method of claim 51, wherein the oligonucleotide is an oligodeoxyribonucleotide or an oligoribonucleotide.

53. The method of claim 42, further comprising repeating said steps (a)-(c) to form an oligonucleotide array.

54. The method of claim 41, wherein the step of aligning the substrate comprises the steps of:

(a) moving the substrate over a marker to establish one or marks on the substrate;

(b) subsequently locating the marks with a camera;

(c) determining misalignment of the substrate relative to the print head with reference to the marks; and

(d) moving the substrate to correct the misalignment.

55. The method of claim 54, wherein the step of moving the substrate to correct the misalignment is done by moving the substrate in a linear motion in X and Y directions and by rotating the substrate.

84. The method of claim 41 or 56, wherein the substrate is porous.

85. The method of claim 50, wherein the biopolymer array is two-dimensional.

86. The method of claim 53 or 57, wherein the oligonucleotide array is two-dimensional.

91. The method of claim 41, wherein said system further comprises a plurality of flow cells.

56. A method of controlling a system for synthesizing oligonucleotides on a substrate, said system capable of processing one or more substrates, using a computer having a memory for storing a control program and data, wherein the system comprises an inkjet print head for spraying a solution comprising a nucleoside or activated nucleoside, on the substrate, a scanning transport for scanning the substrate adjacent to the print head to selectively deposit the solution, an alignment unit for detecting misalignment of the substrate with respect to the print head at each deposition step, a flow cell for treating the substrate, and a treating transport for moving the substrate between the printer head and the flow cell, wherein said treating transport and said scanning transport are different elements, the method comprising the steps of:

(a) placing the substrate with respect to the print head and establishing marks on the substrate;

(b) selectively depositing the solution on the substrate by sending a signal to the print head to spray the solution and by sending a signal to the scanning transport to move the substrate adjacent to the print head so that the solution can be deposited at specified loci on the substrate;

(c) controlling treatment of the substrate by sending a signal to the treating transport to move the substrate to the flow cell and by sending a signal to the flow cell to control operation of the flow cell; and

(d) placing the substrate adjacent to the print head and aligning the substrate with respect to the print head by processing data from the alignment unit and by sending a signal to the scanning transport to move the substrate so as to correct misalignment of the substrate.

57. The method of claim 56, further comprising repeating said steps (a)-(d) to form an oligonucleotide array.

92. The method of claim 56, wherein said system further comprises a plurality of flow cells.

58. An automated system for synthesizing oligonucleotides on a substrate, said automated system capable of processing one or more substrates, comprising:

an inkjet print head having an array of pumps for depositing a nucleoside or activated nucleoside at specified loci on the substrate;

a first translational stage having at least two axes of movement;

a first substrate chuck mounted for movement by the translational stage, the first substrate chuck being adapted to hold the substrate and move adjacent to the print head;

a flow cell that receives the substrate and that exposes the substrate to one or more selected fluids;

a second translational stage that has at least two axes of movement, wherein said first translational stage and said second translational stage are different elements;

a second substrate chuck mounted for movement by the second translational stage, the second substrate chuck being adapted to hold the substrate and move between the print head and the flow cell;

control logic connected to control movement of the first and second translational stages;

a marker positioned adjacent to the print head that can be activated to mark particular loci on the substrate for positionally calibrating the substrate with respect to the print head; and

a camera positioned adjacent to the print head to positionally calibrate the substrate with respect to the print head, wherein the camera is connected to provide images to the control logic.

59. The system of claim 58, wherein the first and the second translational stages are driven by stepping motors.

60. The system of claim 58, wherein the first and second substrate chucks are vacuum chucks.

61. The system of claim 58, wherein the first substrate chuck is rotatable for alignment with the print head.

62. The system of claim 58, further comprising a tip that can be activated to scratch marks at particular loci on the substrate for positionally calibrating the substrate.

63. The system of claim 58, further comprising:

a reservoir connected to the print head to supply the nucleoside or activated nucleoside to the print head, wherein

the reservoir contains said nucleoside or activated nucleoside dissolved in a solvent.

64. The system of claim 63, wherein the solvent has a boiling point of 150°

C. or above, a surface tension of 30 dynes/cm or above, and a viscosity of 0.015 g/(cm)(sec) or above.

65. The system of claim 64, wherein the solvent has the formula (I):

wherein

A=O or S;

X=O, S or N (C₁-C₄ alkyl);

Y=O, S, N(C₂-C₂₀ alkyl) or CH₂; and

R=C₂-C₂₀ straight or branched chain alkyl.

66. The system of claim 65, wherein the solvent is selected from the group consisting of:

N-methyl-2-pyrrolidone;

2-pyrrolidone;

propylene carbonate;

γ

-valerolactone;

6-caprolactam;

ethylene carbonate;

γ

-butyrolactone;

δ

-valerolactone;

1,3-dimethyl-3,4,5,6-tetrahydro-2(1H)-pyrimidinone;

ethylene trithiocarbonate; and

1,3-dimethyl-2-imidazolidinone.

67. The system of claim 64, wherein said solvent is propylene carbonate.

87. The system of claim 58, wherein the substrate is porous.

93. The automated system of claim 58, further comprising a plurality of flow cells.

68. An apparatus programmed for controlling a system for synthesizing a biopolymer on a substrate, said system capable of processing one or more substrates, wherein the system comprises an inkjet print head for spraying a microdroplet on the substrate, a scanning transport for scanning the substrate adjacent to the print head to selectively deposit the microdroplet, an alignment unit for detecting misalignment of the substrate with respect to the print head at each deposition step, a flow cell for treating the substrate, and a treating transport for moving the substrate between the printer head and the flow cell, wherein said treating transport and said scanning transport are different elements, the controller comprising:

(a) means for controlling alignment of the substrate relative to the print head by processing data from the alignment unit and by sending a signal to the scanning transport to move the substrate so as to correct misalignment of the substrate;

(b) means for controlling selective deposition of a microdroplet on the substrate by sending a signal to the print head to spray the microdroplet and by sending a signal to the scanning transport to move the substrate adjacent to the print head so that the microdroplet can be deposited at specified loci on the substrate; and

(c) means for controlling treatment of the substrate by sending a signal to the treating transport to move the substrate to the flow cell and by sending a signal to the flow cell to control operation of the flow cell.

69. The apparatus of claim 68, wherein the inkjet print head contains a solution comprising a monomer unit of a biopolymer.

70. The apparatus of claim 69, wherein the solution further comprises a catalyst.

71. The apparatus of claim 69, wherein the solution comprises a solvent that has a boiling point of 150°

C. or above, a surface tension of 30 dynes/cm or above, and a viscosity of 0.015 g/(cm)(sec) or above.

72. The apparatus of claim 71, wherein the solvent has the formula (I):

wherein

A=O or S;

X=O, S or N (C₁-C₄ alkyl);

Y=O, S, N(C₁-C₄ alkyl) or CH₂; and

R=C₁-C₂₀ straight or branched chain alkyl.

73. The apparatus of claim 68, wherein the solvent is selected from the group consisting of:

N-methyl-2-pyrrolidone;

2-pyrrolidone;

propylene carbonate;

γ

-valerolactone;

6-caprolactam;

ethylene carbonate;

γ

-butyrolactone;

δ

-valerolactone;

1,3-dimethyl- 3,4,5,6 -tetrahydro-2(1H)-pyrimidinone;

ethylene trithiocarbonate; and

1,3-dimethyl-2-imidazolidinone.

88. The apparatus of claim 68 or 75, wherein the substrate is porous.

94. The apparatus of claim 68, wherein said system further comprises a plurality of flow cells.

74. The apparatus of claim wherein said solvent is propylene carbonate.

75. An apparatus programmed for controlling a system for synthesizing a biopolymer on a substrate, said system capable of processing one or more substrates, wherein the system comprises an inkjet print head for spraying a microdroplet on the substrate, a scanning transport for scanning the substrate adjacent to the print head to selectively deposit the microdroplet, an alignment unit for detecting misalignment of the substrate with respect to the print head at each deposition step, a flow cell for treating the substrate, and a treating transport for moving the substrate between the printer head and the flow cell, wherein said treating transport and said scanning transport are different elements, said apparatus comprising one or more computer systems programmed for:

(a) controlling alignment of the substrate relative to the print head by processing data from the alignment unit and by sending a signal to the scanning transport to move the substrate so as to correct misalignment of the substrate;

(b) controlling selective deposition of a microdroplet on the substrate by sending a signal to the print head to spray the microdroplet and by sending a signal to the scanning transport to move the substrate adjacent to the print head so that the microdroplet can be deposited at specified loci on the substrate; and

(c) controlling treatment of the substrate by sending a signal to the treating transport to move the substrate to the flow cell and by sending a signal to the flow cell to control operation of the flow cell.

76. The apparatus of claim 75, further comprising a reservoir connected to the print head to supply a nucleoside or activated nucleoside to the print head, wherein the reservoir contains said nucleoside or activated nucleoside dissolved in a solvent.

77. The apparatus of claim 75, wherein the inkjet print head contains a solution comprising a monomer unit of a biopolymer.

78. The apparatus of claim 77, wherein the solution further comprises a catalyst.

79. The apparatus of claim 77, wherein the solution comprises a solvent that has a boiling point of 150°

C. or above, a surface tension of 30 dynes/cm or above, and a viscosity of 0.015 g/(cm)(sec) or above.

80. The apparatus of claim 79, wherein the solvent has the formula (I):

wherein

A=O or S;

X=O, S or N(C₁-C₄ alkyl);

Y=O, S, N(C₁-C₄ alkyl) or CH₂; and

R=C₁-C₂₀ straight or branched chain alkyl.

81. The apparatus of claim 80, wherein the solvent is selected from the group consisting of:

N-methyl-2-pyrrolidone;

2-pyrrolidone;

propylene carbonate;

γ

-valerolactone;

6-caprolactam;

ethylene carbonate;

γ

-butyrolactone;

δ

-valerolactone;

1,3-dimethyl- 3,4,5,6 -tetrahydro-2(1H)-pyrimidinone;

ethylene trithiocarbonate; and

1,3-dimethyl-2-imidazolidinone.

82. The apparatus of claim 79, wherein said solvent is propylene carbonate.

95. The apparatus of claim 75, wherein said system further comprises a plurality of flow cells.