Oligonucleotide probes and primers comprising universal bases for diagnostic purposes

• US 20030170711A1
• Filed: 02/25/2003
• Published: 09/11/2003
• Est. Priority Date: 10/02/1997
• Status: Active Grant

• Current Assignee: Gen-Probe Incorporated (Hologic Incorporated)
• Original Assignee: Gen-Probe Incorporated (Hologic Incorporated)
• Inventors: Brown, Bob D., Riley, Timothy A.
• Granted Patent: US 8,153,772 B2
• Time in Patent Office: Days
• US Class Current: 435/6
• CPC Class Codes: C07H 21/00 Compounds containing two or more mononucleotide units having separate phosphate or polyphosphate groups linked by saccharide radicals of nucleoside groups, e.g. nucleic acids
• CPC Class Codes: C07H 21/04 with deoxyribosyl as saccharide radical
• CPC Class Codes: C12N 15/1034 Isolating an individual clone by screening libraries
• CPC Class Codes: C12N 15/113 Non-coding nucleic acids modulating the expression of genes, e.g. antisense oligonucleotides; Antisense DNA or RNA; Triplex- forming oligonucleotides; Catalytic nucleic acids, e.g. ribozymes; Nucleic acids used in co-suppression or gene silencing when used in plants C12N15/8218
• CPC Class Codes: C12N 15/1135 against oncogenes or tumor suppressor genes
• CPC Class Codes: C12N 15/1137 against enzymes viral enzymes C12N15/1131; receptors C12N15/1138
• CPC Class Codes: C12N 2310/12 catalytic nucleic acids, e.g. ribozymes
• CPC Class Codes: C12N 2310/31 of the backbone
• CPC Class Codes: C12N 2310/3125 Methylphosphonates
• CPC Class Codes: C12N 2310/313 Phosphorodithioates
• CPC Class Codes: C12N 2310/315 Phosphorothioates
• CPC Class Codes: C12N 2310/318 where the PO2 is completely replaced, e.g. MMI or formacetal
• CPC Class Codes: C12N 2310/3181 Peptide nucleic acid, PNA
• CPC Class Codes: C12N 2310/321 2'-O-R Modification
• CPC Class Codes: C12N 2310/33 of the base
• CPC Class Codes: C12N 2310/346 having a combination of backbone and sugar modifications
• CPC Class Codes: C12N 2310/351 Conjugate
• CPC Class Codes: C12N 2310/3521 Methyl
• CPC Class Codes: C12N 2310/3533 Halogen
• CPC Class Codes: C12Q 1/6883 for diseases caused by alterations of genetic material
• CPC Class Codes: C12Q 2600/156 : Polymorphic or mutational markers

Abstract

Aspects of the invention relate to novel oligonucleotides comprising universal and generic bases for use as primers and probes, as well as, methods of using said oligonucleotides for the diagnosis of disease.

Claims

1. An oligonucleotide comprising between about 10 and 100 bases, wherein at least 2 of said bases are juxtaposed universal bases.

2. The oligonucleotide of claim 1, wherein at least 3 of said bases are juxtaposed universal bases.

3. The oligonucleotide of claim 1, wherein at least 4 of said bases are juxtaposed universal bases.

4. The oligonucleotide of claim 1, wherein at least 5 of said bases are juxtaposed universal bases.

5. The oligonucleotide of claim 1, wherein at least 6 of said bases are juxtaposed universal bases.

6. The oligonucleotide of claim 1, wherein at least 7 of said bases are juxtaposed universal bases.

7. The oligonucleotide of claim 1 wherein said universal bases are selected from the group consisting of:

2-deoxyinosine, 5-nitroindole, 3-nitropyrrole, and 2-deoxynebularine.

8. The oligonucleotide of claim 1, wherein said oligonucleotide comprises between about 10 and about 15 bases.

9. The oligonucleotide of claim 1, wherein said oligonucleotide comprises between about 10 and about 25 bases..

10. The oligonucleotide of claim 1, wherein said oligonucleotide comprises between about 10 and about 50 bases.

11. The oligonucleotide of claim 1, further comprising a non-nucleic acid linker.

12. The oligonucleotide of claim 11, wherein said non-nucleic acid linker is selected from the group consisting of:

Spacer 9, Spacer 18, Spacer C3, and dSpacer.

13. The oligonucleotide of claim 1, wherein said oligonucleotide comprises modifications that decrease the affinity of the oligonucleotide for a target.

14. The oligonucleotide of claim 1, wherein said oligonucleotide comprises modifications that increase the affinity of the oligonucleotide for a target.

15. The oligonucleotide of claim 1, wherein more than about 20% of said bases are universal or generic bases.

16. The oligonucleotide of claim 1, wherein more than about 30% of said bases are universal or generic bases.

19. The oligonucleotide of claim 1 wherein said universal bases are selected from the group consisting of:

2-deoxyinosine, 5-nitroindole, 3-nitropyrrole, and 2-deoxynebularine.

17. An oligonucleotide comprising the formula:

XRY

wherein X comprises between about 5 and 20 modified nucleic acid bases;

wherein R comprises between about 3 and 10 juxtaposed universal bases;

wherein Y comprises between about 5 and about 20 nucleic acid bases;

wherein X, R, and Y are covalently joined; and

wherein more than about 20% of the total number of bases in said oligonucleotide are said juxtaposed universal bases.

18. The oligonucleotide of claim 17, further comprising a non-nucleic acid linker.

20. A method for detecting a polymorphism in a sample of genetic material, comprising:

providing an oligonucleotide comprising between about 10 and 100 bases, wherein at least 2 of said bases are juxtaposed universal bases;

contacting said genetic material with said oligonucleotide; and

identifying whether said oligonucleotide is bound to said genetic material, wherein said universal or generic bases are not located at the site or sites of said polymorphism.

21. The method of claim 20, wherein said mutations are Single Nucleotide Polymorphisms (SNPs).

22. The method of claim 20 further comprising amplifying said mutation in said genetic material.

23. The method of claim 20, wherein said genetic material is DNA.

24. The method of claim 20 wherein said universal bases are selected from the group consisting of:

2-deoxyinosine, 5-nitroindole, 3-nitropyrrole, and 2-deoxynebularine.

25. The method of claim 20, wherein the presence of said polymorphism is diagnostic for a genetic disease.

26. A method for diagnosing a genetic disease in a patient, comprising:

providing an oligonucleotide specific for a DNA sequence related to said disease, wherein said oligonucleotide comprises between about 10 and 100 bases, and wherein at least 2 of said bases are juxtaposed universal bases;

contacting genetic material from said patient with said oligonucleotide; and

identifying whether said oligonucleotide is bound to said genetic material, wherein binding of said oligonucleotide to said genetic material is indicative of said genetic disease.

27. The method of claim 26, wherein said genetic disease comprises a DNA sequence comprising a Single Nucleotide Polymorphisms (SNPs).

28. The method of claim 26 further comprising amplifying said DNA sequence.

29. The method of claim 26, wherein said genetic material is DNA.

30. The method of claim 26 wherein said universal bases are selected from the group consisting of:

2-deoxyinosine, 5-nitroindole, 3-nitropyrrole, and 2-deoxynebularine.

31. The method of claim 26, wherein said oligonucleotide comprises 10 to 50 bases.

32. The method of claim 26, wherein said oligonucleotide comprises 10 to 25 bases.

33. The method of claim 26, wherein said oligonucleotide comprises 10 to 15 bases.