COMPOSITIONS FOR DETECTION AND ANALYSIS OF POLYNUCLEOTIDES USING LIGHT HARVESTING MULTICHROMOPHORES

• US 20080064042A1
• Filed: 09/12/2007
• Published: 03/13/2008
• Est. Priority Date: 08/26/2002
• Status: Active Grant

• Current Assignee: Brent Gaylord, Guillermo Bazan, Shu Wang
• Original Assignee: Brent Gaylord, Guillermo Bazan, Shu Wang
• Inventors: Gaylord, Brent, Wang, Shu, Bazan, Guillermo
• Granted Patent: US 7,629,448 B2
• Time in Patent Office: Days
• US Class Current: 435/6
• CPC Class Codes: C12Q 1/6813 Hybridisation assays
• CPC Class Codes: C12Q 1/6818 involving interaction of two or more labels, e.g. resonant energy transfer
• CPC Class Codes: C12Q 1/6825 Nucleic acid detection involving sensors
• CPC Class Codes: C12Q 1/6834 Enzymatic or biochemical coupling of nucleic acids to a solid phase
• CPC Class Codes: C12Q 2523/313 Irradiation, e.g. UV irradiation
• CPC Class Codes: C12Q 2563/107 fluorescence
• CPC Class Codes: C12Q 2565/107 Alteration in the property of hybridised versus free label oligonucleotides
• CPC Class Codes: Y10T 436/143333 Saccharide [e.g., DNA, etc.]

Abstract

Methods, compositions and articles of manufacture for assaying a sample for a target polynucleotide are provided. A sample suspected of containing the target polynucleotide is contacted with a polycationic multichromophore and a sensor polynucleotide complementary to the target polynucleotide. The sensor polynucleotide comprises a signaling chromophore to receive energy from the excited multichromophore and increase emission in the presence of the target polynucleotide. The methods can be used in multiplex form. Kits comprising reagents for performing such methods are also provided.

Claims

1. A polynucleotide sensing solution comprising:

a signaling chromophore; and

a polycationic multichromophore that is capable of transferring energy to the signaling chromophore upon excitation when brought into proximity thereto, wherein a greater amount of energy can be produced from the signaling chromophore in the presence of the polynucleotide being sensed when the multichromophore is excited.

2. The polynucleotide sensing solution of claim 1, further comprising a second signaling chromophore that can absorb energy from the signaling chromophore in the presence of the target polynucleotide being sensed and emit light.

3. The polynucleotide sensing solution of claim 2, wherein the second signaling chromophore is a polynucleotide-specific dye.

4. The polynucleotide sensing solution of claim 1, further comprising a sensor polynucleotide that is single-stranded and is complementary to the target polynucleotide being sensed.

5. The polynucleotide sensing solution of claim 1, wherein the signaling chromophore is a polynucleotide-specific dye that can absorb energy from the multichromophore in the presence of the target polynucleotide being sensed and emit light.

6. The polynucleotide sensing solution of claim 1, wherein the signaling chromophore is a polynucleotide-specific dye that can absorb energy from the multichromophore in the presence of the target polynucleotide being sensed and transfer energy to a second signaling chromophore conjugated to a sensor polynucleotide which then emits light.

7. The polynucleotide sensing solution of claim 1, wherein the multichromophore comprises a structure that is selected from the group consisting of a saturated polymer, a conjugated polymer, an aggregate of conjugated molecules, a dendrimer, and a semiconductor nanocrystal.

8. The polynucleotide sensing solution of claim 7, wherein the multichromophore comprises a dendrimer.

9. The polynucleotide sensing solution of claim 7, wherein the multichromophore comprises a conjugated polymer.

10. The polynucleotide sensing solution of claim 9, wherein the conjugated polymer has the structure

11. The polynucleotide sensing solution of claim 7, wherein the multichromophore comprises an aggregate of conjugated molecules.

12. The polynucleotide sensing solution of claim 11, wherein the aggregate comprises molecules having the structure

13. The polynucleotide sensing solution of claim 4, wherein the solution further comprises a sufficient amount of an organic solvent to decrease hydrophobic interactions between the sensor polynucleotide and the multichromophore.

14. The polynucleotide sensing solution of claim 1, wherein the solution comprises a plurality of different sensor polynucleotides having corresponding different sequences, said different sensor polynucleotides comprising a corresponding different signaling chromophore, wherein each of said different sensor polynucleotides can selectively hybridize to a corresponding different target polynucleotide.

15. The polynucleotide sensing solution of claim 1, wherein the signaling chromophore is a fluorophore.

16. The polynucleotide sensing solution of claim 15, wherein the fluorophore is selected from a semiconductor nanocrystal, a fluorescent dye, and a lanthanide chelate.

17. A sensing complex comprising:

a sensor polynucleotide that is single-stranded and is complementary to a target polynucleotide, said sensor polynucleotide attached to a signaling chromophore; and

a polycationic multichromophore complexed with the sensor polynucleotide, wherein the polycationic multichromophore is capable of transferring energy to the signaling chromophore upon excitation when in proximity thereto, wherein the complex can emit light from the signaling chromophore upon excitation of the multichromophore in the absence of target polynucleotide, and wherein a greater amount of emitted light is produced from the signaling chromophore upon excitation of the multichromophore in the presence of target polynucleotide.

18. A kit for assaying a sample for a target polynucleotide comprising:

a sensor polynucleotide that is single-stranded and is complementary to the target polynucleotide;

a signaling chromophore;

a polycationic multichromophore that is capable of transferring energy to the signaling chromophore upon excitation when brought into proximity thereto, wherein said sensor polynucleotide interacts with the multichromophore and a detectibly greater amount of emitted light is produced from the signaling chromophore upon excitation of the multichromophore in the presence of target polynucleotide; and

a housing for retaining the components of the kit.

19. The kit of claim 18, further comprising a second signaling chromophore that can absorb energy from the signaling chromophore in the presence of target and emit light.

20. The kit of claim 19, wherein the second signaling chromophore is a polynucleotide-specific dye.

21. The kit of claim 18, further comprising instructions provided with said housing that describe how to use the components of the kit to assay the sample for the target polynucleotide.

22. The kit of claim 18, wherein the multichromophore comprises a structure that is selected from the group consisting of a saturated polymer, a conjugated polymer, an aggregate of conjugated molecules, a dendrimer, and a semiconductor nanocrystal.

23. The kit of claim 22, wherein the multichromophore comprises a conjugated polymer.

24. The kit of claim 18, wherein the kit further comprises an organic solvent capable of decreasing hydrophobic interactions between the sensor polynucleotide and the multichromophore.

25. The kit of claim 18, wherein the signaling chromophore is a fluorophore selected from a semiconductor nanocrystal, a fluorescent dye, and a lanthanide chelate.