Detection of Nucleic Acids Through Amplification of Surrogate Nucleic Acids

• US 20120009577A1
• Filed: 08/10/2011
• Published: 01/12/2012
• Est. Priority Date: 11/10/2005
• Status: Active Grant

• Current Assignee: Affymetrix, Inc. (Thermo Fisher Scientific Incorporated)
• Original Assignee: Affymetrix, Inc. (Thermo Fisher Scientific Incorporated)
• Inventors: Luo, Yuling, Bui, Son
• Granted Patent: US 8,288,522 B2
• Time in Patent Office: Days
• US Class Current: 435/6.11
• CPC Class Codes: C12Q 1/6844 Nucleic acid amplification reactions
• CPC Class Codes: C12Q 1/686 Polymerase chain reaction [PCR]
• CPC Class Codes: C12Q 2537/143 Multiplexing, i.e. use of multiple primers or probes in a single reaction, usually for simultaneously analyse of multiple analysis
• CPC Class Codes: C12Q 2561/113 Real time assay

Abstract

Methods for detecting and optionally quantitating one or more target nucleic acids are provided, in which a surrogate nucleic acid is captured to each target nucleic acid, amplified, and detected. Compositions, kit, and systems related to the methods are also described.

Claims

1-26. -26. (canceled)

27. A composition comprising:

a first target nucleic acid;

a nucleic acid comprising a first surrogate nucleic acid;

a first set of one or more surrogate capture probes, each of which surrogate capture probes is capable of hybridizing simultaneously to the first target nucleic acid and to the nucleic acid comprising the first surrogate nucleic acid; and

one or more primers for amplifying the first surrogate nucleic acid.

28. The composition of claim 27, wherein the nucleic acid comprising the first surrogate nucleic acid is hybridized to the first set of surrogate capture probes, which surrogate capture probes are hybridized to the first target nucleic acid, whereby the first surrogate nucleic acid is physically associated with the first target nucleic acid at a molar ratio of about 1:

1.

29. The composition of claim 27, wherein the nucleic acid comprising the first surrogate nucleic acid is hybridized to the first set of surrogate capture probes, which surrogate capture probes are hybridized to the first target nucleic acid, whereby the first surrogate nucleic acid is physically associated with the first target nucleic acid at a molar ratio of at least about 2:

1, at least about 3;

1, at least about 5;

1, or at least about 10;

1 first surrogate nucleic acid;

first target nucleic acid.

30. The composition of claim 27, wherein one or more copies of the nucleic acid comprising the first surrogate nucleic acid are associated with a copy of the first target nucleic acid;

wherein the first set of surrogate capture probes comprises a subset of surrogate capture probes for each of the one or more copies of the nucleic acid comprising the first surrogate nucleic acid; and

wherein each subset of surrogate capture probes comprises n surrogate capture probes, where n is at least two.

31. The composition of claim 27, wherein the first set of surrogate capture probes comprises two or more surrogate capture probes.

32. The composition of claim 31, wherein the surrogate capture probes hybridize to nonoverlapping polynucleotide sequences in the first target nucleic acid.

33. The composition of claim 27, comprising a solid support.

34. The composition of claim 33, wherein the solid support comprises a plurality of particles.

35. The composition of claim 33, comprising a first set of m target capture probes, where m is at least one, which target capture probes are capable of hybridizing to the first target nucleic acid.

36. The composition of claim 35, wherein the target capture probes hybridize to nonoverlapping polynucleotide sequences in the first target nucleic acid.

37. The composition of claim 35, wherein m is at least two.

38. The composition of claim 35, wherein the solid support comprises a first support capture probe bound to the solid support, and wherein the target capture probes of the first set are capable of hybridizing simultaneously to the first target nucleic acid and to the first support capture probe.

39. The composition of claim 27, comprising amplified first surrogate nucleic acid.

40. The composition of claim 39, comprising one or more reagents for detecting the amplified first surrogate nucleic acid.

42. The composition of claim 27, wherein the first surrogate nucleic acid is a DNA.

45. The composition of claim 27, comprising a second nucleic acid comprising a second surrogate nucleic acid;

a second set of one or more surrogate capture probes, each of which surrogate capture probes is capable of hybridizing simultaneously to a second target nucleic acid and to the second nucleic acid comprising the second surrogate nucleic acid; and

the second target nucleic acid and/or one or more primers for amplifying the second surrogate nucleic acid.

50. The composition of claim 30, wherein the 5′

end of each of the surrogate capture probes in the first set hybridizes to the first target nucleic acid while the 3′

end hybridizes to the nucleic acid comprising the first surrogate nucleic acid, or wherein the 3′

end of each of the surrogate capture probes in the first set hybridizes to the first target nucleic acid while the 5′

end hybridizes to the nucleic acid comprising the first surrogate nucleic acid.

51. The composition of claim 27, wherein the first surrogate nucleic acid is physically associated with the first target nucleic acid.

41. (canceled)

43-44. -44. (canceled)

46. A kit comprising:

a nucleic acid comprising a first surrogate nucleic acid;

a first set of one or more surrogate capture probes, each of which surrogate capture probes is capable of hybridizing simultaneously to a first target nucleic acid and to the nucleic acid comprising the first surrogate nucleic acid;

a solid support comprising a first support capture probe bound to the solid support;

a first set of m target capture probes, where m is at least one, which target capture probes are capable of hybridizing simultaneously to the first target nucleic acid and to the first support capture probe; and

one or more primers for amplifying the first surrogate nucleic acid;

packaged in one or more containers.

49. The kit of claim 46, further comprising:

a second nucleic acid comprising a second surrogate nucleic acid;

a second set of one or more surrogate capture probes, each of which surrogate capture probes is capable of hybridizing simultaneously to a second target nucleic acid and to the second nucleic acid comprising the second surrogate nucleic acid; and

a second set of m target capture probes, where m is at least one, which target capture probes are capable of hybridizing simultaneously to the second target nucleic acid and to the first support capture probe.

47-48. -48. (canceled)