Methods for generating databases and databases for identifying polymorphic genetic markers

• US 8,229,677 B2
• Filed: 12/21/2009
• Issued: 07/24/2012
• Est. Priority Date: 10/13/1999
• Status: Expired due to Fees

• Current Assignee: Sequenom Incorporated (Laboratory Corporation Of America Holdings)
• Original Assignee: Sequenom Incorporated (Laboratory Corporation Of America Holdings)
• Inventors: Braun, Andreas, Koster, Hubert, Yip, Ping, Rodi, Charles, He, Liyan, Chiu, Norman, Van den Boom, Dirk, Jurinke, Christiane
• Primary Examiner(s): Clow, Lori A
• Application Number:
  US12/643,933
• Publication Number:
  US 20100292930A1
• Time in Patent Office: 946 Days
• Field of Search: None
• US Class Current: 702/19
• CPC Class Codes: C12Q 1/6827 for detection of mutation or polymorphism
• CPC Class Codes: C12Q 1/6872 involving mass spectrometry
• CPC Class Codes: C12Q 1/6883 for diseases caused by alterations of genetic material
• CPC Class Codes: C12Q 2521/301 Endonuclease
• CPC Class Codes: C12Q 2535/125 Allele specific primer extension
• CPC Class Codes: C12Q 2561/125 Ligase Detection Reaction [LDR]
• CPC Class Codes: C12Q 2565/627 being a mass spectrometer
• CPC Class Codes: C12Q 2600/156 Polymorphic or mutational markers
• CPC Class Codes: Y10T 436/24 Nuclear magnetic resonance, electron spin resonance or other spin effects or mass spectrometry

Abstract

Processes and methods for creating a database of genomic samples from healthy human donors, methods that use the database to identify and correlate polymorphic genetic markers and other markers with diseases and conditions are provided.

Claims

1. A method for determining whether a polymorphism correlates with a gene or pathway involved in the onset and progression of disease comprising:

a) identifying a polymorphism in a first ethnic group, wherein a disease or biochemical pathway is known to occur in high frequency in the first ethnic group;

b) pooling samples obtained from healthy individuals from said first ethnic group, and pooling samples obtained from healthy individuals from a second ethnic group, wherein healthy individuals are selected only on the basis of being healthy, thereby providing pooled samples;

c) determining the frequency of the polymorphism in said pooled samples,

d) determining a difference in frequency of the polymorphism in healthy individuals between said ethnic groups, wherein the polymorphism has not been correlated with a disease; and

e) associating the polymorphism with the disease or a biochemical pathway that occurs with high frequency in the first ethnic group.

2. The method of claim 1, wherein the frequency of said polymorphism in said samples is in a database, and said database is sorted to identify correlations between the frequency of said polymorphism and said first ethnic group or said second ethnic group.

3. The method of claim 1, wherein said polymorphism comprises a SNP.

4. The method of claim 1, further comprising identifying the locus of said polymorphism and assessing or deducing the function of a gene at said locus.

5. The method of claim 1, wherein said sample comprises body tissue or fluid from said individual.

6. The method of claim 1, wherein said sample comprises DNA.

7. The method of claim 1, wherein the frequency of the polymorphism is determined using mass spectrometry.

8. The method of claim 7, wherein the mass spectrometric format is selected from among Matrix Assisted Laser Desorption/lonization, Time of Flight (MALDI TOF), Electrospray (ES), IR-MALDI, Ion Cyclotron Resonance (ICR), Fourier Transform, and combinations thereof.

9. The method of claim 1, wherein said method comprises obtaining genomic nucleic acid from a sample from a healthy individual.

10. The method of claim 1, further comprising amplifying a portion of the genomic nucleic acid to produce amplified fragments thereof.

11. The method of claim 1, wherein the polymorphism is identified by a method comprising primer oligo base extension.

12. The method of claim 11, wherein the primer oligo base extension comprises hybridizing a nucleic acid molecule from a sample from a healthy individual with a primer oligonucleotide that is complementary to the nucleic acid molecule at a site adjacent to the polymorphic marker.

13. The method of claim 12, further comprising:

f) optionally immobilizing the nucleic acid molecule onto a solid support, to produce an immobilized nucleic acid molecule;

g) contacting the optionally-immobilized nucleic acid molecule with a composition comprising a dideoxynucleoside triphosphate or a 3′

-deoxynucleoside triphosphate and a polymerase, so that only a dideoxynucleoside or 3′

-deoxynucleoside triphosphate that is complementary to the polymorphic marker is extended onto the primer; and

h) detecting the extended primer, thereby identifying the polymorphism.

14. The method of claim 1, wherein the polymorphism is identified by a method comprising:

identifying samples by sorting a database comprising datapoints representative of a plurality of individuals from whom biological samples are obtained, wherein each datapoint is associated with data representative of the organism type and other identifying information, wherein said database is sorted according to a selected parameter to identify samples that match the selected parameter;

isolating a nucleic acid molecule from each identified sample;

pooling each isolated nucleic acid molecule; and

identifying the polymorphism in the nucleic acid molecule by a method comprising primer oligo base extension.

15. The method of claim 1, wherein the polymorphism is identified by a method comprising:

identifying samples by sorting a database comprising datapoints representative of a plurality of individuals from whom biological samples are obtained, wherein each datapoint is associated with data representative of the organism type and other identifying information, wherein said database is sorted according to a selected parameter to identify samples that match the selected parameter;

isolating a biopolymer from each identified sample;

pooling each isolated biopolymer;

cleaving the pooled biopolymers to produce fragments thereof;

obtaining a mass spectrum of the resulting fragments and comparing the mass spectrum with a control mass spectrum to identify differences between the spectra and thereby identifying any polymorphisms;

wherein;

the control mass spectrum is obtained from either

samples represented by datapoints in said database that were not selected by sorting said database;

or

samples identified by sorting said database according to a different selected parameter.

16. The method of claim 1, wherein the polymorphism is identified by a method comprising:

isolating a biopolymer from samples of body tissue or fluid from a plurality of individuals of an ethnic group;

pooling each isolated biopolymer;

cleaving the pooled biopolymers to produce fragments thereof;

obtaining a mass spectrum of the resulting fragments;

determining a frequency of each fragment, whereby an average frequency is calculated; and

comparing the frequency of each fragment to identify fragments present in amounts lower than the average frequency, thereby identifying the polymorphism.

17. The method of claim 15, wherein the biopolymers comprise genomic nucleic acid molecules.