Method and apparatus for synthesis of arrays of DNA probes

• US 6,375,903 B1
• Filed: 02/22/1999
• Issued: 04/23/2002
• Est. Priority Date: 02/23/1998
• Status: Expired due to Term

• Current Assignee: Wisconsin Alumni Research Foundation (University of Wisconsin System)
• Original Assignee: Wisconsin Alumni Research Foundation (University of Wisconsin System)
• Inventors: Green, Roland, Sussman, Michael R., Singh-Gasson, Sangeet, Cerrina, Francesco, Blattner, Frederick R.
• Primary Examiner(s): Marschel, Ardin H.
• Application Number:
  US09/253,460
• Time in Patent Office: 1,156 Days
• Field of Search: 422/50, 422/62, 422/68.1, 422/69, 422/129, 422/131, 422/134, 422/149, 422/186, 422/188, 422/189, 422/190, 422/193, 422/211, 422/219, 422/232, 435/6, 435/283.1, 435/286.1, 435/286.2, 435/286.4, 435/286.5, 435/289.1, 435/292.1, 435/299.1, 435/305.1, 536/25.3, 530/333, 530/334
• US Class Current: 422/131
• 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 2219/00439 using micromirror arrays
• CPC Class Codes: B01J 2219/00527 Sheets
• CPC Class Codes: B01J 2219/00529 DNA chips
• 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 2219/00608 DNA chips
• CPC Class Codes: B01J 2219/00612 the surface being inorganic
• CPC Class Codes: B01J 2219/00626 Covalent
• CPC Class Codes: B01J 2219/00637 by coating it with another layer
• CPC Class Codes: B01J 2219/00641 the porous medium being continuous, e.g. porous oxide substrates
• CPC Class Codes: B01J 2219/00659 Two-dimensional arrays
• CPC Class Codes: B01J 2219/00689 using computers
• CPC Class Codes: B01J 2219/00711 Light-directed synthesis
• CPC Class Codes: B01J 2219/00722 Nucleotides
• CPC Class Codes: B82Y 30/00 Nanotechnology for materials or surface science, e.g. nanocomposites
• CPC Class Codes: C07B 2200/11 Compounds covalently bound to a solid support
• 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: C40B 40/00 : Libraries per se, e.g. arrays, mixtures
• CPC Class Codes: C40B 40/06 : Libraries containing nucleotides or polynucleotides, or derivatives thereof
• CPC Class Codes: C40B 60/14 : for creating libraries
• CPC Class Codes: G02B 26/0841 : the reflecting element being moved or deformed by electrostatic means
• CPC Class Codes: G03F 7/2002 : with visible light or UV light, through an original having an opaque pattern on a transparent support, e.g. film printing, projection printing; by reflection of visible or UV light from an original such as a printed image

Abstract

The synthesis of arrays of DNA probes sequences, polypeptides, and the like is carried out using a patterning process on an active surface of a substrate. An image is projected onto the active surface of the substrate utilizing an image former that includes a light source that provides light to a micromirror device comprising an array of electronically addressable micromirrors, each of which can be selectively tilted between one of at least two positions. Projection optics receives the light reflected from the micromirrors along an optical axis and precisely images the micromirrors onto the active surface of the substrate, which may be used to activate the surface of the substrate. The first level of bases may then be applied to the substrate, followed by development steps, and subsequent exposure of the substrate utilizing a different pattern of micromirrors, with further repeats until the elements of a two dimensional array on the substrate surface have an appropriate base bound thereto. The micromirror array can be controlled in conjunction with a DNA synthesizer supplying appropriate reagents to a flow cell containing the active substrate to control the sequencing of images presented by the micromirror array in coordination of the reagents provided to the substrate.

Claims

1. Apparatus for use in synthesis of arrays of DNA probes, comprising:

(a) a substrate with an active surface on which the arrays may be formed;

(b) a flow cell enclosing the active surface of the substrate and having ports for applying DNA synthesis reagents into the flow cell which can be flowed over the active surface of the substrate; and

(c) an image former providing a high precision, two-dimensional light image projected onto the substrate active surface, comprising;

(1) a light source providing a light beam;

(2) a micromirror device receiving the light beam from the source and formed of an array of electronically addressable micromirrors, each of which can be selectively tilted between one of at least two separate positions, wherein in one of the positions of each micromirror the light from the source incident upon the micromirror is deflected away from an optical axis and in a second of the at least two positions of the micromirror the light is reflected along the optical axis; and

(3) projection optics receiving the light reflected from the micromirrors along the optical axis and imaging the pattern of the micromirrors onto the active surface of the substrate, wherein the light is directed from the micromirror onto the active surface by reflective optical elements.

2. The apparatus of claim 1 wherein the micromirror device is formed of a two dimensional array of micromirrors.

3. The apparatus of claim 1 including a lens for collimating the beam from the light source to provide a collimated beam projected onto the micromirror array at an oblique angle to a main optical axis that extends from the micromirror array to the substrate, and wherein in one position of each micromirror the light is reflected along the optical axis through the projection optics to the substrate and in a second position of each micromirror the light from the source is reflected at an angle off the main axis of the projection system and away from the substrate.

4. The apparatus of claim 1 further including a filter receiving the light from the source and which selectively passes only desired wavelengths through to the micromirror array.

5. The apparatus of claim 1 wherein the substrate is transparent and light from the image former is passed through the transparent substrate to be imaged on the active surface of the substrate which is opposite to the surface which initially receives the light from the image former.

6. The apparatus of claim 1 further including a computer connected to the micromirror device to provide command signals to control the deflection of the mirrors in the micromirror array to provide a desired pattern for projection onto the substrate.

7. The apparatus of claim 1 wherein the light provided by the light source is in the range of ultraviolet to near ultraviolet wavelengths.

8. The apparatus of claim 7 including a filter receiving the light from the source which selectively passes wavelengths in the ultraviolet and near ultraviolet and blocks longer wavelengths including infrared.

9. The apparatus of claim 8 wherein the filter includes a dichroic mirror that reflects the selected wavelengths and passes the wavelengths to be blocked.

10. The apparatus of claim 1 wherein the pattern of the micromirrors that is imaged onto the active surface of the substrate is reduced in size with respect to the size of the array of micromirrors.

11. The apparatus of claim 1 wherein the projection optics is telecentric.

12. The apparatus of claim 1 wherein the reflective optical elements include a concave mirror and a convex mirror, the concave mirror reflecting light from the micromirror device to the convex mirror which reflects it back to the concave mirror which reflects the light to the substrate where it is imaged.

13. The apparatus of claim 12 including a planar mirror that reflects the light from the concave mirror to the substrate.

14. The apparatus of claim 1 including a flow cell having a housing composed of a lower base and upper cover section and a gasket mounted on the base, wherein the substrate is a transparent glass slide secured between the upper cover section and the base to define a sealed reaction chamber between the substrate and the base that is sealed by the gasket, and channels extending through the housing from the input port to the reaction chamber and from the reaction chamber to the output port, the active surface of the substrate facing the sealed reaction chamber.

15. The apparatus of claim 14 including means for detachably securing the substrate between the lower base and upper cover section to allow the substrate to be replaced.

30. The apparatus of claim 1 further including a DNA synthesizer connected to supply reagents to the flow cell.

16. Apparatus for use in synthesis arrays of DNA probes, comprising:

(a) a substrate with an active surface on which the arrays may be formed;

(b) a flow cell enclosing the active surface of the substrate and having ports for applying DNA synthesis reagents into the flow cell which can be flowed over the active surface of the substrate; and

(c) an image former providing a high precision, two-dimensional light image projected onto the substrate active surface, comprising;

(1) a light source providing a light beam;

(2) a micromirror device receiving the light beam from the source and formed of an array of electronically addressable micromirrors, wherein the micromirror device is formed of a two dimensional array of micromirrors, cach of which can be selectively tilted between one of at least two separate positions, wherein in one of the positions of each micromirror the light from the source incident upon the micromirror is deflected away from an optical axis and in a second of the at least two positions of the micromirror the light is reflected along the optical axis; and

(3) projection optics receiving the light reflected from the micromirrors along the optical axis and imaging the pattern of the micromirrors onto the active surface of the substrate, wherein the projection optics is telecetric and wherein light is directed from the micromirrors to the active surface is by reflective optical elements.

17. The apparatus of claim 16 including a lens for collimating the beam from the light source to provide a collimated beam projected onto the micromirror array at an oblique angle to a main optical axis that extends from the micromirror array to the substrate, and wherein in one position of each micromirror the light is reflected along the optical axis through the projection optics to the substrate and in a second position of each micromirror the light from the source is reflected at an angle off the main axis of the projection system and away from the substrate.

18. The apparatus of claim 16 further including a filter receiving the light from the source and which selectively passes only desired wavelengths through to the micromirror array.

19. The apparatus of claim 16 wherein the substrate is transparent and light from the image former is passed through the transparent substrate to be imaged on the active surface of the substrate which is opposite to the surface which initially receives the light from the image former.

20. The apparatus of claim 16 further including a computer connected to the micromirror device to provide command signals to control the deflection of the mirrors in the micromirror array to provide a desired pattern for projection onto the substrate.

21. The apparatus of claim 16 wherein the light provided by the light source is in the range of ultraviolet to near ultraviolet wavelengths.

22. The apparatus of claim 21 including a filter receiving the light from the source which selectively passes wavelengths in the ultraviolet and near ultraviolet and blocks longer wavelengths including infrared.

23. The apparatus of claim 22 wherein the filter includes a dichroic mirror that reflects the selected wavelengths and passes the wavelengths to be blocked.

24. The apparatus of claim 16 wherein the pattern of the micromirrors that is imaged onto the active surface of the substrate is reduced in size with respect to the size of the array of micromirrors.

25. The apparatus of claim 16 including refractive lenses between the light source and the micromirror device that form a Kohler illumination system.

26. The apparatus of claim 16 wherein the reflective optical elements include a concave mirror and a convex mirror, the concave mirror reflecting light from the micromirror device to the convex mirror which reflects it back to the concave mirror which reflects the light to the substrate where it is imaged.

27. The apparatus of claim 26 including a planar mirror that reflects the light from the concave mirror to the substrate.

28. The apparatus of claim 16 wherein the flow cell has a housing composed of a lower base and upper cover section is a gasket mounted on the base, wherein the substrate is a transparent glass slide secured between the upper cover section and the base to define a sealed reaction chamber between the substrate and the base that is sealed by the gasket, and channels extending through the housing from the input port to the reaction chamber and from the reaction chamber to the output port, the active surface of the substrate facing the sealed reaction chamber.

29. The apparatus of claim 28 including means for detachably securing the substrate between the lower base and upper cover section to allow the substrate to be replaced.

31. The apparatus of claim 16 further including a DNA synthesizer connected to supply reagents to the flow cell.