Methods and devices based upon a novel form of nucleic acid duplex on a surface

• US 7,354,710 B2
• Filed: 07/11/2002
• Issued: 04/08/2008
• Est. Priority Date: 07/11/2001
• Status: Expired due to Term

• Current Assignee: Baylor College of Medicine (Baylor College), Genomics USA, Inc.
• Original Assignee: Genomics USA, Inc.
• Inventors: Powdrill, Tom, Belosludtsev, Yuri, Hogan, Michael, Mitra, Rahul, Lemeshko, Sergy
• Primary Examiner(s): Kim; Young J.
• Application Number:
  US10/193,938
• Publication Number:
  US 20030134299A1
• Time in Patent Office: 2,098 Days
• Field of Search: 435/6, 536/24.3, 536/24.33
• 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 2219/00376 in multiple or parallel arrangements
• CPC Class Codes: B01J 2219/00387 Applications using probes
• CPC Class Codes: B01J 2219/00497 Features relating to the solid phase supports
• CPC Class Codes: B01J 2219/005 Beads
• CPC Class Codes: B01J 2219/00529 DNA chips
• CPC Class Codes: B01J 2219/00576 fluorophore
• 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 2219/00653 the compounds being bound to electrodes embedded in or on the solid supports
• CPC Class Codes: B01J 2219/00659 Two-dimensional arrays
• CPC Class Codes: B01J 2219/00677 Ex-situ synthesis followed by deposition on the substrate
• CPC Class Codes: B01J 2219/00691 using robots
• CPC Class Codes: B01J 2219/00722 Nucleotides
• CPC Class Codes: C12Q 1/6837 using probe arrays or probe chips C12Q1/6874 takes precedence
• CPC Class Codes: C12Q 2523/308 Adsorption or desorption
• CPC Class Codes: C40B 60/14 for creating libraries

Abstract

The present invention relates to simple method to fabricate DNA hybridization devices based upon adsorptive attachment of oligonucleotides to a positively charged surface. Such adsorbed oligonucleotide probes form a densely packed monolayer, which retains capacity for base-pair specific hybridization with a solution state nucleic acid target strand to form the duplex. However, both strand dissociation kinetics and the rate of DNase digestion suggest on symmetry grounds that solution-state nucleic acid binds to such adsorbed oligonucleotides to form a highly asymmetric and unwound duplex, with structural details that are substantially different from that known for the Watson-Crick DNA duplex. This novel nucleic acid duplex form can serve as the basis for a new class of hybridization device and methods for their use. It is also disclosed that new methods of nucleic acid duplex detection can be developed which are based upon the interaction of enzymes and dye labels with the unique structural characteristics of the non-helical duplex described herein. Preferred implementations of the invention include DNA microarrays, bead-based nucleic acid analysis, microelectronic devices to detect nucleic acid hybridization and more traditional methods of laboratory analysis, including hybridization on membranous and other solid supports.

Claims

1. A biomolecular hybridization device comprising:

a substrate having an aminosilanized surface permanently and covalently attached thereto; and

an adsorbed monolayer formed from an about twice saturating amount of unmodified single-stranded oligonucleotides all of which are about 12 to 16 bases in length adsorbed to the aminosilanized surface as a saturated film of constrained oligonucleotides on the surface via direct non-covalent phosphate-surface adsorptive contact of substantially all phosphate groups of each oligonucleotide at a density of one phosphate group per about 1.5 square nanometers of surface, wherein each constrained oligonucleotide base plane is presented from the surface in a manner effective to dissociably hybridize to a complementary single-stranded nucleic acid with asymmetric, substantially non-helical base pairing without alteration of the oligonucleotide base plane presentation and without oligonucleotide phosphate group dissociation from the surface.

2. The biomolecular hybridization device of claim 1 wherein said device further comprises a single-stranded nucleic acid reversibly hybridized to the oligonucleotides as a substantially non-helical duplex of 12 to 16 base pairs long adsorbed to said, surface.

3. The biomolecular hybridization device of claim 1 wherein the form of said substrate is a slide or a bead.

4. The biomolecular hybridization device of claim 2 wherein the nucleic acid is DNA.

5. The biomolecular hybridization device of claim 2 wherein the nucleic acid is RNA.

6. The biomolecular hybridization device of claim 1 further comprising a capping material disposed on said surface.

7. The method biomolecular hybridization device of claim 6 wherein said capping material is comprises deposition of a surfactant or an acid anhydride.