Methods and compositions for analyzing nucleic acid molecules utilizing sizing techniques

• US 7,052,846 B2
• Filed: 06/19/2003
• Issued: 05/30/2006
• Est. Priority Date: 01/23/1996
• Status: Expired due to Fees

• Current Assignee: Agilent Technologies, Inc.
• Original Assignee: Operon Biotechnologies, Inc. (Eurofins Scientific SE)
• Inventors: Van Ness, Jeffrey, Tabone, John C, Mulligan, John T, Howbert, J Jeffry
• Primary Examiner(s): Riley, Jezia
• Application Number:
  US10/465,524
• Publication Number:
  US 20060057566A1
• Time in Patent Office: 1,076 Days
• Field of Search: 435/6, 435/91.1, 435/91.2, 536/24.3, 536/24.33
• US Class Current: 435/6.12
• CPC Class Codes: C07B 2200/11 Compounds covalently bound to a solid support
• 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: C12Q 1/6825 Nucleic acid detection involving sensors
• CPC Class Codes: C12Q 1/6846 Common amplification features
• CPC Class Codes: C12Q 1/6872 involving mass spectrometry
• CPC Class Codes: C12Q 2525/191 incorporating an adaptor
• CPC Class Codes: C12Q 2563/113 the label being electroactive, e.g. redox labels
• CPC Class Codes: C12Q 2563/149 Particles, e.g. beads
• CPC Class Codes: C12Q 2563/167 Mass label
• CPC Class Codes: C40B 40/00 Libraries per se, e.g. arrays, mixtures

Abstract

Tags and linkers specifically designed for a wide variety of nucleic acid reactions are disclosed, which are suitable for a wide variety of nucleic acid reactions wherein separation of nucleic acid molecules based upon size is required.

Claims

1. A kit for genotyping, comprising a plurality of amplification primer pairs, wherein at least one of the primers has the formula:

T^ms-L-X

wherein,

T^ms is an organic group detectable by mass spectrometry, comprising carbon, at least one of hydrogen and fluoride, and optional atoms selected from oxygen, nitrogen, sulfur, phosphorus and iodine;

L is an organic group which allows a T^ms-containing moiety to be cleaved from the remainder of the compound, wherein the T^ms-containing moiety comprises a functional group which supports a single ionized charge state when the compound is subjected to mass spectrometry and is selected from tertiary amine, quaternary amine and organic acid; and

X is a nucleic acid fragment wherein L is conjugated to X at a location other than the 3′

end of X; and

each primer pair associates with a different locus;

the compound also having the formula;

2. The kit according to claim 1 wherein the plurality is at least 3.

3. A kit according to claim 1 wherein the plurality is at least 5.

4. A kit for genotyping, comprising a plurality of amplification primer pairs, wherein at least one of the primers has the formula:

T^ms-L-X

wherein,

T^ms is an organic group detectable by mass spectrometry, comprising carbon, at least one of hydrogen and fluoride, and optional atoms selected from oxygen, nitrogen, sulfur, phosphorus and iodine;

L is an organic group which allows a T^ms-containing moiety to be cleaved from the remainder of the compound, wherein the T^ms-containing moiety comprises a functional group which supports a single ionized charge state when the compound is subjected to mass spectrometry and is selected from tertiary amine, quaternary amine and organic acid; and

X is a nucleic acid fragment wherein L is conjugated to X at a location other than the 3′

end of X; and

each primer pair associates with a different locus;

the compound also having the formula;

5. The kit according to claim 4 wherein the -Amide-T⁵ is selected from:

6. The kit according to claim 4 wherein -Amide-T⁵ is selected from:

7. The kit according to claim 4 wherein the plurality is at least 3.

8. The kit according to claim 4 wherein the plurality is at least 5.

9. A kit for genotyping, comprising a plurality of amplification primer pairs, wherein at least one of the primers has the formula:

T^ms-L-X

wherein,

T^ms is an organic group detectable by mass spectrometry, comprising carbon, at least one of hydrogen and fluoride, and optional atoms selected from oxygen, nitrogen, sulfur, phosphorus and iodine;

L is an organic group which allows a T^ms-containing moiety to be cleaved from the remainder of the compound, wherein the T^ms-containing moiety comprises a functional group which supports a single ionized charge state when the compound is subjected to mass spectrometry and is selected from tertiary amine, quaternary amine and organic acid; and

X is a nucleic acid fragment wherein L is conjugated to X at a location other than the 3′

end of X; and

each primer pair associates with a different locus;

the compound also having the formula;

10. The kit according to claim 9 wherein the -Amide-T⁵ is selected from:

11. The kit according to claim 9 wherein -Amide-T⁵ is selected from:

12. The kit according to claim 9 wherein the plurality is at least 3.

13. The kit according to claim 9 wherein the plurality is at least 5.