Methods and devices for nucleic acid sequence determination

• US 7,635,562 B2
• Filed: 05/25/2005
• Issued: 12/22/2009
• Est. Priority Date: 05/25/2004
• Status: Expired due to Fees

• Current Assignee: Fluidigm Corporation
• Original Assignee: Helicos Biosciences Corporation
• Inventors: Harris, Timothy, Buzby, Philip Richard, DiMeo, James Joseph, Jarosz, Mirna, Gill, Jaime
• Primary Examiner(s): Martinell; James
• Application Number:
  US11/137,928
• Publication Number:
  US 20060019276A1
• Time in Patent Office: 1,672 Days
• Field of Search: None
• US Class Current: 435/6.11
• 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 2219/00432 Photolithographic masks
• CPC Class Codes: B01J 2219/00436 Maskless processes
• CPC Class Codes: B01J 2219/00497 Features relating to the solid phase supports
• CPC Class Codes: B01J 2219/005 Beads
• CPC Class Codes: B01J 2219/00511 Walls of reactor vessels
• CPC Class Codes: B01J 2219/00527 Sheets
• CPC Class Codes: B01J 2219/00574 radioactive
• CPC Class Codes: B01J 2219/00576 fluorophore
• CPC Class Codes: B01J 2219/00585 Parallel 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 2219/0061 The surface being organic
• CPC Class Codes: B01J 2219/00612 the surface being inorganic
• CPC Class Codes: B01J 2219/00617 by chemical means
• CPC Class Codes: B01J 2219/00619 using hydrophilic or hydrophobic regions
• CPC Class Codes: B01J 2219/00621 by physical means, e.g. trenches, raised areas
• CPC Class Codes: B01J 2219/00626 Covalent
• CPC Class Codes: B01J 2219/00628 Ionic
• CPC Class Codes: B01J 2219/0063 Other, e.g. van der Waals forces, hydrogen bonding
• 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 2219/00675 : In-situ synthesis on the substrate
• CPC Class Codes: B01J 2219/00677 : Ex-situ synthesis followed by deposition on the substrate
• CPC Class Codes: B01J 2219/00689 : using computers
• CPC Class Codes: B01J 2219/00707 : separated from the reactor apparatus
• 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: C12Q 1/68 : involving nucleic acids
• CPC Class Codes: C12Q 1/6837 : using probe arrays or probe chips C12Q1/6874 takes precedence
• CPC Class Codes: C12Q 1/6874 : involving nucleic acid arrays, e.g. sequencing by hybridisation
• CPC Class Codes: C12Q 2549/125 : using sterilising/blocking agents, e.g. albumin

Abstract

Methods of the invention comprise methods and devices for nucleic acid sequence determination. Generally, the invention relates to preparing a substrate for sequencing a target nucleic acid.

Claims

1. A method for conducting a chemical reaction on a surface, the method comprising the steps of:

attaching a plurality of first nucleic acid molecules via covalent linkages to reactive chemical groups on a surface;

blocking the surface using a blocking solution comprising potassium phosphate;

conducting a chemical reaction between said first nucleic acid molecules and second reactant molecules comprising an optically detectable label wherein said conducting step comprises exposing said nucleic acid to a primer under conditions sufficient to extend said nucleic acid by at least one base;

rinsing unincorporated optically labeled nucleotides from the surface; and

observing individually optically resolvable labels incorporated in the one or more first nucleic acids on the surface.

2. The method of claim 1, wherein said first nucleic acids are DNA or RNA.

3. The method of claim 1, wherein said optically detectable label is a fluorescent molecule.

4. The method of claim 3, wherein said fluorescent molecule is selected from the group consisting of Cyanine-3 and Cyanine-5.

5. The method of claim 1, wherein the reactive chemical groups are epoxide groups.

6. The method of claim 1, wherein the first nucleic acid molecules are attached to the reactive chemical groups indirectly by means of a biotin-streptavidin binding pair wherein biotin is covalently attached to the surface.

7. The method of claim 1, further comprising the step of exposing the surface to a drying agent.

8. The method of claim 7, wherein said drying agent is PRB, EtOH, air or N₂.

9. The method of claim 1, further comprising determining nucleic acid sequences of one or more of the first nucleic acid molecules.

10. The method of claim 9, wherein said determining step comprises conducting sequencing by synthesis.

11. The method of claim 1, wherein the surface comprises a silica coating containing exposed epoxide groups.

12. The method as in claim 1, wherein pH of the blocking solution is 8.5.

13. A method for conducting a chemical reaction on a surface, the method comprising the steps of:

attaching a plurality of first nucleic acid molecules via covalent linkages to reactive chemical groups on an analytical surface;

blocking the surface using a blocking solution comprising potassium phosphate;

conducting a chemical reaction using second reactant molecules comprising an optically detectable label thereby incorporating said second reactant molecules;

rinsing unincorporated optically labeled second reactant molecules from the surface; and

observing individually optically resolvable labels on the surface.