Informative nucleic acid arrays and methods for making same

• US 6,635,423 B2
• Filed: 01/16/2001
• Issued: 10/21/2003
• Est. Priority Date: 01/14/2000
• Status: Expired due to Fees

• Current Assignee: Thomas P. Dooley
• Original Assignee: Integriderm, Inc.
• Inventors: Curto, Ernest V., Davis, Richard L. Jr., Dooley, Thomas P.
• Primary Examiner(s): Horlick, Kenneth R.
• Assistant Examiner(s): STRZELECKA, TERESA E
• Application Number:
  US09/759,377
• Publication Number:
  US 20010046671A1
• Time in Patent Office: 1,008 Days
• Field of Search: 435/6, 435/968, 435/973, 536/24.3, 436/63, 702/19
• US Class Current: 435/6.14
• CPC Class Codes: C12Q 1/6837 using probe arrays or probe chips C12Q1/6874 takes precedence
• CPC Class Codes: C12Q 1/6876 Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes
• CPC Class Codes: C12Q 2539/101 Subtraction analysis
• CPC Class Codes: C12Q 2565/515 characterised by the interaction between or sequential use of two or more arrays
• CPC Class Codes: C12Q 2600/158 Expression markers
• CPC Class Codes: Y10S 435/968 High energy substrates, e.g. fluorescent, chemiluminescent, radioactive

Abstract

The present invention is directed to methods for making and using informative nucleic acid arrays (e.g., DNA including cDNA, RNA, PNA) for research and other applications in various disciplines or areas of interest. Examples of such disciplines include, without limitation, dermatology, pharmacology, toxicology, oncology, gynecology, urology, gastroenterology, as well as studies of sentinel gene discovery, signature gene discovery, mechanism of action, drug screening, drug metabolism, etc. The informative nucleic acid arrays of the present invention may contain only the gene sequences that are of interest in a particular area of interest or application, and may exclude other gene sequences.

Claims

1. A method for identifying a plurality of genes that are differentially expressed between dissimilar biological samples for use in an informative array, comprising:

providing a first set of heterogeneous nucleic acid probes derived from a first biological sample;

providing a second set of heterogeneous nucleic acid probes derived from a second biological sample;

hybridizing a nucleic acid array comprising a plurality of sequences derived from genes of a biological process with the first set of probes and determining a first level of expression for sequences of the array;

hybridizing the array with said second set of probes and determining a second level of expression for sequences of the array;

identifying a plurality of genes that are differentially expressed in said biological process by comparing the first level of expression with said second level of expression for hybridized sequences; and

establishing a ranking of the identified genes by a step selected from the group of steps consisting of;

determining an absolute value of the difference between the first level of expression and the second level of expression, and ranking genes having a higher difference over genes having a lower difference; and

determining a standard deviation of the difference between the first level of expression and the second level of expression, and ranking genes having a higher standard deviation over genes having a lower standard deviation.

2. The method of claim 1, wherein the biological process is selected from the group consisting of the processes of absorption, distribution, metabolism, and excretion of drugs, toxins and chemicals.

3. The method of claim 1, wherein the first and second biological samples are selected from the group consisting of samples of skin, skin appendages, oral tissue, gastrointestinal tissue, neural tissue, renal tissue, hepatic tissue, urogenital tissue and combinations thereof.

4. The method of claim 1, wherein the first and second biological samples are selected from the group consisting of samples of cells, cell cultures, cell lines, tissues, organs, organelles, biological fluids, whole organisms and combinations thereof.

5. The method of claim 1, wherein the differentially expressed genes are derived from genes of epithelial cells or tissues.

6. The method of claim 1, wherein the differentially expressed genes are derived from genes of cells selected from the group consisting of skin, skin appendages, oral tissue, gastrointestinal tissue, neural tissue, renal tissue, hepatic tissue, urogenital tissue and combinations thereof.

7. The method of claim 1, wherein the step of identifying comprises:

quantifying the difference in the first level of expression and the second level of expression for hybridized sequences by applying an algorithm to the first level of expression and the second level of expression, performing cluster analysis on the first level of expression and the second level of expression, or performing dendrogram analysis on the first level of expression and the second level of expression.

8. The method of claim 1, further comprising the step of hybridizing the array with three or more sets of probes, each derived from a different biological sample, and determining a level of expression with regard to hybridization to sequences of the array.

9. The method of claim 1, wherein the step of establishing a ranking comprises:

determining an absolute value for the difference between the first level of expression and the second level of expression for the identified genes, and ranking genes having a higher difference over genes having a lower difference.

10. The method of claim 1, wherein the step of establishing a ranking comprises:

determining a standard deviation of the difference between the first level of expression and the second level of expression and ranking genes having a higher standard deviation over genes having a lower standard deviation.

11. The method of claim 9, wherein the step of establishing a ranking of the identified genes that are differentially expressed comprises:

ranking the identified genes with a bias against more highly or more lowly expressed genes.

12. The method of claim 10, wherein the step of establishing a ranking of the identified genes that are differentially expressed comprises:

ranking the identified genes with a bias against more highly or more lowly expressed genes.

13. A method for selecting a plurality of genes for an informative nucleic acid array for a biological process, comprising:

identifying genes that are differentially expressed in a biological process;

establishing a ranking of the differentially expressed genes by a step selected from the group of steps consisting of;

determining an absolute value of the difference between the first level of expression and the second level of expression, and ranking genes having a higher difference over genes having a lower difference; and

determining a standard deviation of the difference between the first level of expression and the second level of expression, and ranking genes having a higher standard deviation over genes having a lower standard deviation; and

placing sequences derived from ranked differentially expressed genes on the informative array.

14. The method of claim 13, wherein the genes are expressed in the biological process.

15. The method of claim 13, wherein the step of identifying comprises:

providing a first set of nucleic acid probes derived from a first biological sample;

providing a second set of nucleic acid probes derived from a second biological sample, wherein said second biological sample is dissimilar from said first biological sample;

hybridizing the first set of probes to the array and determining the first level of expression for hybridized genes;

hybridizing the second set of probes to the array and determining the second level of expression for hybridized genes; and

identifying genes that are differentially expressed by comparing the first level of expression with the second level of expression for hybridized genes.

16. The method of claim 13, wherein the step of identifying comprises:

quantifying the difference in the first level of expression and the second level of expression for hybridized genes by applying an algorithm to the first level of expression and the second level of expression, performing cluster analysis on the first level of expression and the second level of expression, or performing dendrogram analysis on the first level of expression and the second level of expression.

17. The method of claim 13, further comprising applying the selected genes to a filter surface, a glass surface, a plastic surface, a solid bead, or a gene chip surface.

18. The method of claim 13, wherein the step of establishing a ranking comprises employing at least one analytical method to rank the differentially expressed genes.

19. The method of claim 13, further comprising the step of:

using the informative nucleic acid array to perform gene expression analyses on RNA isolated from cells, cell cultures, cell lines, tissues, organs, organelles, biological fluids or whole organisms.

20. The method of claim 13, further comprising the step of:

using the array to identify at least one signature biomarker gene, wherein the signature biomarker gene is expressed in a cell-specific manner, a differentiation-specific manner, or a tissue-specific manner.

21. The method of claim 13, further comprising the step of:

using the informative nucleic acid array to identify at least one sentinel biomarker gene, wherein the sentinel biomarker gene is expressed in response to treatment by an agent which comprises a drug, a toxin, a chemical, a temperature, a pressure, or electromagnetic radiation.

23. The method of claim 13, wherein the step of establishing a ranking comprises:

determining an absolute value for the difference between the first level of expression and the second level of expression for the identified genes, and ranking genes having a higher difference over genes having a lower difference.

24. The method of claim 13, wherein the step of establishing a ranking comprises:

determining a standard deviation of the difference between the first level of expression and the second level of expression and ranking genes having a higher standard deviation over genes having a lower standard deviation.

25. The method of claim 23, wherein the step of establishing a ranking of the identified genes that are differentially expressed comprises:

ranking the identified genes with a bias against more highly or more lowly expressed genes.

26. The method of claim 24, wherein the step of establishing a ranking of the identified genes that are differentially expressed comprises:

ranking the identified genes with a bias against more highly or more lowly expressed genes.

22. A method for converting a nucleic acid array into an informative array comprising:

providing a first set of heterogeneous nucleic acid probes derived from a first biological sample;

providing a different, second set of heterogeneous nucleic acid probes derived from a second biological sample;

hybridizing a nucleic acid array comprising a plurality of sequences with the first set of probes and determining a first level of expression for sequences of the array;

hybridizing the array with said second set of probes and determining a second level of expression for sequences of the array;

identifying a plurality of genes that are differentially expressed in said biological process based on a difference between the first level of expression and the second level of expression for identified genes, by a step selected from the group of steps consisting of;

determining an absolute value for the difference between the first level of expression and the second level of expression, and ranking genes having a higher difference over genes having a lower difference; and

determining a standard deviation of the difference between the first level of expression and the second level of expression, and ranking genes having a higher standard deviation over genes having a lower standard deviation; and

selecting genes from the plurality of identified differentially expressed genes for inclusion on the informative array.

27. The method of claim 22, wherein the step of establishing a ranking comprises:

determining an absolute value for the difference between the first level of expression and the second level of expression for the identified genes, and ranking genes having a higher difference over genes having a lower difference.

28. The method of claim 22, wherein the step of establishing a ranking comprises:

determining a standard deviation of the difference between the first level of expression and the second level of expression and ranking genes having a higher standard deviation over genes having a lower standard deviation.

29. The method of claim 27, wherein the step of establishing a ranking of the identified genes that are differentially expressed comprises:

ranking the identified genes with a bias against more highly or more lowly expressed genes.

30. The method of claim 28, wherein the step of establishing a ranking of the identified genes that are differentially expressed comprises:

ranking the identified genes with a bias against more highly or more lowly expressed genes.