METHODS OF DIRECT GENOMIC SELECTION USING HIGH DENSITY OLIGONUCLEOTIDE MICROARRAYS

• US 20100093986A1
• Filed: 02/04/2008
• Published: 04/15/2010
• Est. Priority Date: 02/02/2007
• Status: Abandoned Application

• Current Assignee: Emory University
• Original Assignee: Emory University
• Inventors: Zwick, Michael E., Okou, David T.
• Application Number:
  US12/524,252
• Publication Number:
  US 20100093986A1
• Time in Patent Office: Days
• US Class Current: 536/23.1
• CPC Class Codes: C12Q 1/6837 using probe arrays or probe chips C12Q1/6874 takes precedence

Abstract

The present disclosure encompasses methods (hereinafter termed ‘Microarray-based Genomic Selection’ (MGS), capable of isolating user-defined unique genomic sequences from complex eukaryotic genomes.

Claims

1. A method of isolating user-defined unique gene sequences from complex eukaryotic genomes comprising:

isolating genomic from a human or animal;

shearing the genomic DNA into fragments;

repairing the genomic DNA fragments;

ligating adapters to the genomic DNA fragments;

hybridizing the genomic DNA fragments to oligonucleotides of interest of a high density long oligonucleotide microarray;

eluting of the genomic DNA fragments bound to oligonucleotides of interest on the microarray; and

amplifying the eluted DNA fragments.

2. The method of claim 1, further comprising resequencing of the eluted DNA fragments.

3. The method of claim 1, wherein the shearing is physical shearing.

4. The method of claim 3, wherein the shearing is selected from sonication, nebulization, or a combination thereof.

5. The method of claim 1, wherein repairing includes using blunt end formation or the addition of 3′

-A extensions to the genomic DNA fragments.

6. The method of claim 1, wherein repairing the genomic DNA fragments includes the addition of 3′

-A extensions to the genomic DNA fragments.

7. The method of claim 1, wherein the adapters do not substantially self ligate, are unique relative to the DNA genome, and are complimentary to one another.

8. The method of claim 1, wherein the adaptors have the nucleotide sequences according to SEQ ID NOs:

1 and 2.

9. A method of isolating user-defined unique gene sequences from complex eukaryotic genomes comprising:

isolating genomic from a human or animal;

shearing the genomic DNA into fragments, wherein the shearing is physical shearing selected from sonication, nebulization, or a combination thereof;

repairing the genomic DNA fragment, wherein repairing the genomic DNA fragments includes the addition of 3′

-A extensions to the genomic DNA fragments;

ligating a plurality of adapters to the genomic DNA fragments, wherein the adaptors are blunt-end ligated to the genomic DNA fragments, and wherein the adapters have a 3′

-T extension, do not substantially self ligate, are unique relative to the DNA genome, and are complimentary to one another, and wherein the adaptors have the nucleotide sequences according to SEQ ID NOs;

1 and 2;

hybridizing the genomic DNA fragments to oligonucleotides of interest of a high density long oligonucleotide microarray;

eluting of the genomic DNA fragments bound to oligonucleotides of interest on the microarray;

amplifying the eluted DNA fragments; and

resequencing of the eluted DNA fragments.