Identification of molecular sequence signatures and methods involving the same

• US 8,084,197 B2
• Filed: 05/08/2007
• Issued: 12/27/2011
• Est. Priority Date: 09/19/1996
• Status: Expired due to Fees

• Current Assignee: Affymetrix, Inc. (Thermo Fisher Scientific Incorporated)
• Original Assignee: Affymetrix, Inc. (Thermo Fisher Scientific Incorporated)
• Inventors: Lockhart, David J., Wong, Gordon G., Langer-Safer, Pennina
• Primary Examiner(s): Whisenant, Ethan C
• Application Number:
  US11/745,987
• Publication Number:
  US 20090029865A1
• Time in Patent Office: 1,694 Days
• Field of Search: 435/6, 536/22.1, 536/23.1, 536/24.3
• US Class Current: 435/6
• CPC Class Codes: B01J 2219/00608 DNA chips
• 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 2219/00626 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 2219/00702 Processes involving means for analysing and characterising the products
• 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: C12Q 1/6827 for detection of mutation or polymorphism
• CPC Class Codes: C12Q 1/6837 using probe arrays or probe chips C12Q1/6874 takes precedence
• CPC Class Codes: C12Q 2565/515 characterised by the interaction between or sequential use of two or more arrays
• CPC Class Codes: C40B 40/06 Libraries containing nucleotides or polynucleotides, or derivatives thereof

Abstract

Novel means and methods for analyzing hybridization data derived from hybridization assays between a target nucleic acid and differently sequenced polynucleotide probes involve selecting probe sets that define reference sequences for sequence signatures and deriving useful data about the nature of the target nucleic acid molecule based on its hybridization to the probes. The methods are useful for determining whether the target contains a nucleic acid or polypeptide sequence signature, whether the target encodes a member of a gene family, or whether the target is derived from one of any number of genes.

Claims

1. A method for determining whether a nucleic acid sample from a patient contains nucleic acid encoding a signature sequence characteristic of a pathogen said method comprising:

obtaining a nucleic acid containing sample from said patient;

providing a probe array comprising a first plurality of polynucleotide probe sets each probe set defining a different reference nucleotide sequence wherein each reference nucleotide sequence signature is between 15 and 30 bases in length and codes for a different amino acid sequence that is a signature sequence characteristic of said pathogen; and

a second plurality of polynucleotide probe sets each probe set defining a degenerate set of nucleotide sequences encoding each different amino acid sequence and wherein the probe set defining said degenerate set comprises at least 25% of the nucleotide sequences encoding said degenerate set;

hybridizing said nucleic acid sample to said probe array to generate a hybridization pattern; and

analyzing said hybridization pattern to determine if nucleic acid encoding a sequence signature characteristic of said pathogen is present in the sample.

2. The method of claim 1, wherein the probe set defining said degenerate set of nucleotide sequences comprises at least 50% of the degenerate set of nucleotide sequences encoding each different amino acid sequence.

3. The method of claim 1, wherein the probe set defining said degenerate set of nucleotide sequences comprises at least 75% of the degenerate set of nucleotide sequences encoding each different amino acid sequence.

4. The method of claim 1, wherein the probe set defining said degenerate set of nucleotide sequences comprises at least 95% of the degenerate set of nucleotide sequences encoding each different amino acid sequence.

5. The method of claim 1, wherein the pathogen is a mycobacterium.

6. A method for determining whether a sample from a human patient has a sequence from a member of a gene family characteristic of a pathogen comprising:

obtaining a sample comprising nucleic acid from said human patient;

providing a polynucleotide probe array comprising, for each of at least two different members of the gene family, a set of polynucleotide probes that define a reference nucleotide sequence from the member of the gene family;

hybridizing said sample comprising nucleic acid to said probe array to generate a hybridization pattern comprising hybridization data; and

processing the hybridization data to determine if the reference nucleotide sequence from one of the members of the gene family is present in the nucleic acid sample.

7. The method of claim 6 wherein the step of processing is performed by a programmable digital computer.

8. The method of claim 6 wherein the polynucleotide probe array further comprises, for each of the members of the gene family, a probe set defining a highly conserved region of the gene and a probe set defining a highly variable region of the gene.

9. The method of claim 6 wherein the polynucleotide probe array further comprises, for each of the members of the gene family, probe sets defining at least two highly conserved regions of the gene and probe sets defining at least two highly variable regions of the gene.

10. The method of claim 6 wherein the reference nucleotide sequence codes for an amino acid sequence and the array further comprises probe sets defining the degenerate set of nucleotide sequences encoding the amino acid sequence.

11. The method of claim 6, wherein the reference nucleotide sequence is a highly conserved region of the gene family.

12. The method of claim 6, wherein said reference nucleotide sequence is between 9 and 100 nucleotides in length.

13. The method of claim 6 wherein the probe array has a density of at least four hundred different polynucleotides sequences per square centimeter.

14. The method of claim 6 wherein the pathogen is a mycobacterium.