COMPOSITIONS FOR DETECTION AND ANALYSIS OF POLYNUCLEOTIDES USING LIGHT HARVESTING MULTICHROMOPHORES

• US 20130190193A1
• Filed: 01/23/2012
• Published: 07/25/2013
• Est. Priority Date: 06/20/2002
• Status: Active Grant

• Current Assignee: Brent S. Gaylord, Guillermo C. Bazan
• Original Assignee: Regents of the University of California (University of California)
• Inventors: Bazan, Guillermo C., Gaylord, Brent S.
• Granted Patent: US 9,371,559 B2
• Time in Patent Office: Days
• US Class Current: 506/9
• CPC Class Codes: B82Y 15/00 Nanotechnology for interacting, sensing or actuating, e.g. quantum dots as markers in protein assays or molecular motors
• CPC Class Codes: C12Q 1/6818 involving interaction of two or more labels, e.g. resonant energy transfer
• CPC Class Codes: C12Q 2525/107 incorporating a peptide nucleic acid
• CPC Class Codes: C12Q 2527/125 Specific component of sample, medium or buffer
• CPC Class Codes: C12Q 2565/107 Alteration in the property of hybridised versus free label oligonucleotides

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 PNA complementary to the target polynucleotide. The sensor PNA comprises a signaling chromophore to absorb energy from the excited multichromophore and emit light 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

1-19. -19. (canceled)

20. A composition comprising:

a water soluble conjugated polymer, and

a signaling chromophore,

wherein the composition is in an aqueous solution, and

wherein a greater amount of energy is emitted from the signaling chromophore upon excitation of the conjugated polymer than from direct excitation of the signaling chromophore.

21. The composition of claim 20, wherein the aqueous solution comprises a biological sample.

22. The composition of claim 21, wherein the biological sample comprises a target polynucleotide.

23. The composition of claim 22, wherein the target polynucleotide is not amplified.

24. The composition of claim 22, wherein the target is a polynucleotide produced via an amplification reaction.

25. The composition of claim 22, wherein the target polynucleotide is selected from the group consisting of mRNA, rRNA, tRNA, hnRNA, ssRNA, ssDNA viral genomes, genomic DNA, mitochondrial DNA, chloroplast DNA, dsRNA or dsDNA viral genomes, plasmids, phage, and viroids.

26. The composition of claim 20, wherein the conjugated polymer transfers energy to the signaling chromophore.

27. The composition of claim 26, wherein the conjugated polymer transfers energy to the signaling chromophore by a mechanism selected from the group consisting of resonant energy transfer, fluorescence resonance energy transfer (FRET), and quantum charge exchange.

28. The composition of claim 26, wherein the energy transfer mechanism is short range.

29. The energy transfer complex of claim 20, wherein light emission from the signaling chromophore is 8 times greater than the amount of light obtained upon direct excitation of the signaling chromophore.

30. The energy transfer complex of claim 20, wherein light emission from the signaling chromophore is 25 times greater than the amount of light obtained upon direct excitation of the signaling chromophore.

31. An energy transfer complex comprising:

a polynucleotide-binding sensor molecule,

a water-soluble conjugated polymer, and

a signaling chromophore capable of receiving energy from an excited state of the conjugated polymer.

32. The energy transfer complex of claim 31, wherein light emission from the signaling chromophore is greater than the amount of light obtained upon direct excitation of the signaling chromophore.

33. The energy transfer complex of claim 31, wherein light emission from the signaling chromophore is 8 times greater than the amount of light obtained upon direct excitation of the signaling chromophore.

34. The energy transfer complex of claim 31, wherein light emission from the signaling chromophore is 25 times greater than the amount of light obtained upon direct excitation of the signaling chromophore.

35. The energy transfer complex of claim 31, wherein the emission from the signaling chromophore is enhanced by using higher molecular weight water soluble conjugated polymers.

36. The energy transfer complex of claim 31, wherein different signaling chromophores are used to generate different emission signals simultaneously.

37. The energy transfer complex of claim 31, wherein the fluorophore is selected from the group consisting of a semiconductor nanocrystal, a fluorescent dye, a green fluorescent protein and a lanthanide chelate.

38. The energy transfer complex of claim 31, wherein the fluorophore is a fluorescent dye.

39. The energy transfer complex of claim 31, wherein the complex is incorporated onto a substrate.

40. The energy transfer complex of claim 31, wherein the complex is in an aqueous solution.

41. The energy transfer complex of claim 38, further comprising a buffer.