Method of identifying virtual representations of nucleotide sequences

• US 8,694,263 B2
• Filed: 05/21/2004
• Issued: 04/08/2014
• Est. Priority Date: 05/23/2003
• Status: Active Grant

• Current Assignee: Cold Spring Harbor Laboratory
• Original Assignee: Cold Spring Harbor Laboratory
• Inventors: Healy, John, Lucito, Robert, Wigler, Michael H
• Primary Examiner(s): Skibinsky, Anna
• Application Number:
  US10/851,779
• Publication Number:
  US 20050032095A1
• Time in Patent Office: 3,609 Days
• Field of Search: None
• US Class Current: 702/19
• CPC Class Codes: C07H 21/04 with deoxyribosyl as saccharide radical

Abstract

The invention provides oligonucleotide probes that can be used to hybridize to a representation of nucleic acid sequences. Compositions containing the probes such as microarrays are also provided. The invention also provides methods of using these probes and compositions in therapeutic, diagnostic, and research applications. Systems and methods for using a word counting algorithm that can quickly and accurately count the number of times a particular string of characters (i.e., nucleotides) appears in a nucleotide sequence (e.g., a genome) are provided. This algorithm can be used to identify the oligonucleotide probes of the invention. The algorithm uses a transform of a genome and an auxiliary data structure to count the number of times a particular word occurs in the genome.

Claims

1. A method of identifying an oligonucleotide, the method comprising

(A) cleaving a genome of at least Z basepairs in silico with a restriction enzyme to generate a plurality of predicted nucleic acid molecules,

(B) generating a virtual representation of said genome by identifying predicted nucleic acid molecules, wherein each predicted nucleic acid molecule has a length of 200-1,200 basepairs, inclusive;

(C) calculating the following;

(i) Z≧

1×

10⁸;

(ii) 300≧

K≧

30;

(iii) the integer closest to (log₄(Z)+2)≧

L₁≧

the integer closest to log₄(Z);

(iv) X is the integer closest to D₁×

(K−

L₁+1);

(v) Y is the integer closest to D₂×

(K−

L₁+1);

(vi) 1.5≧

D₁≧

1; and

(vii) 1>

D₂≧

0.5;

(D) selecting oligonucleotides each having a length of K nucleotides, inclusive, and each with at least 90% sequence identity to a predicted nucleic acid molecule in (B);

(E) identifying all of the L₁-mers occurring in each oligonucleotide; and

(F) selecting one or more oligonucleotides that have a sum total value of L₁-mer counts in the virtual representation of no fewer than Y and no more than X, wherein an L₁-mer is a subregion of the oligonucleotide having a length of L₁ nucleotides, wherein an L₁-mer count is the number of times the sequence represented by one L₁-mer occurs in the genome, and wherein the sum total value of L₁-mer counts is the sum of every L₁-mer count of the oligonucleotide occurring in the virtual representation.

2. The method of claim 1, wherein the oligonucleotide is a nucleic acid probe.

3. The method of claim 1, wherein K is 40 to 70.

4. The method of claim 1, wherein the virtual representation has no more than R % of the complexity of said genome, wherein 70%≧

R %≧

1%.

5. The method of claim 4, wherein R % is 1 to 2.5%.

6. The method of claim 1, wherein Z is at least 1×

10⁹.

7. The method of claim 1, wherein the genome is a mammalian genome.

8. The method of claim 1, wherein the genome is a human genome.

9. The method of claim 1, wherein D₁ is 1.

10. The method of claim 1, wherein D₂ is 0.5.

11. The method of claim 1, wherein L₁ is 15, 16, 17, 18, 19, 20, 21, 22, 23 or 24.

12. The method of claim 1, wherein said representation is obtained with two or more different restriction endonucleases.