Parallel or antiparallel, homologous or complementary binding of nucleic acids or analogues thereof to form duplex, triplex or quadruplex complexes

• US 20050014140A1
• Filed: 11/12/2002
• Published: 01/20/2005
• Est. Priority Date: 12/21/1999
• Status: Active Grant

• Current Assignee: Ingeneus Inc.
• Original Assignee: Ingeneus Inc.
• Inventors: Kandic, Ivana, Erikson, Glen H., Picard, Pierre, Daksis, Jasmine I.
• Granted Patent: US 7,309,569 B2
• Time in Patent Office: Days
• US Class Current: 435/6
• CPC Class Codes: C12Q 1/6816 characterised by the detection means C12Q1/6804 takes precedence
• CPC Class Codes: C12Q 1/6827 for detection of mutation or polymorphism
• CPC Class Codes: C12Q 1/6839 Triple helix formation or other higher order conformations in hybridisation assays
• CPC Class Codes: C12Q 2525/101 incorporating non-naturally occurring nucleotides, e.g. inosine
• CPC Class Codes: C12Q 2525/113 incorporating modified backbone
• CPC Class Codes: C12Q 2527/125 Specific component of sample, medium or buffer
• CPC Class Codes: C12Q 2537/119 Triple helix formation
• CPC Class Codes: C12Q 2563/107 fluorescence
• CPC Class Codes: C12Q 2563/137 Metal/ion, e.g. metal label
• CPC Class Codes: C12Q 2563/173 staining/intercalating agent, e.g. ethidium bromide

Abstract

A complex includes: (1) a probe containing a heteropolymeric probe sequence of nucleic acids or nucleic acid analogues; and (2) a target containing a heteropolymeric target sequence of nucleic acids or nucleic acid analogues, wherein: (a) at least one of the probe and the target is purified or synthetic; and (b) the heteropolymeric probe sequence is bonded to the heteropolymeric target sequence by Watson-Crick complementary base interaction or by homologous base interaction, provided that when the complex is a duplex and the heteropolymeric probe sequence is antiparallel to the heteropolymeric target sequence, the heteropolymeric probe sequence is bonded to the heteropolymeric target sequence by homologous base interaction, and provided that when the complex is a triplex, the complex is free of recombination proteins. A method for assaying a target includes detecting formation of the complex.

Claims

1. A complex comprising:

a probe containing a heteropolymeric probe sequence of nucleic acids or nucleic acid analogues; and

a target containing a heteropolymeric target sequence of nucleic acids or nucleic acid analogues,

wherein;

(a) at least one of said probe and said target is purified or synthetic; and

(b) said heteropolymeric probe sequence is bonded to said heteropolymeric target sequence by Watson-Crick complementary base interaction or by homologous base interaction, provided that when said complex is a duplex and said heteropolymeric probe sequence is antiparallel to said heteropolymeric target sequence, said heteropolymeric probe sequence is bonded to said heteropolymeric target sequence by homologous base interaction, and provided that when said complex is a triplex, said complex is free of recombination proteins.

2. The complex of claim 1, wherein said complex is a duplex and said heteropolymeric probe sequence is bound to said heteropolymeric target sequence by homologous base interaction or by Watson-Crick complementary base interaction and said heteropolymeric probe sequence is parallel or antiparallel to said heteropolymeric target sequence.

3. The complex of claim 1, wherein one of said probe and said target is single-stranded, the other of said probe and said target is double-stranded, and said complex is a triplex.

4. The complex of claim 3, wherein said complex is free of PNA.

5. The complex of claim 3, wherein said heteropolymeric probe sequence and said heteropolymeric target sequence have parallel directionality, and said heteropolymeric probe sequence is bonded to said heteropolymeric target sequence by homologous base interaction.

6. The complex of claim 3, wherein said heteropolymeric probe sequence and said heteropolymeric target sequence have parallel directionality, and said heteropolymeric probe sequence is bonded to said heteropolymeric target sequence by Watson-Crick complementary base interaction.

7. The complex of claim 3, wherein:

(a) said heteropolymeric probe sequence and said heteropolymeric target sequence have parallel directionality;

(b) said target includes a first strand containing said heteropolymeric target sequence and a second strand containing a second heteropolymeric target sequence complementary and antiparallel to said first heteropolymeric target sequence; and

(c) said heteropolymeric probe sequence is bonded to said heteropolymeric target sequence by homologous base interaction and said heteropolymeric probe sequence is bonded to said second heteropolymeric target sequence by Watson-Crick complementary base interaction.

8. The complex of claim 3, wherein said heteropolymeric probe sequence and said heteropolymeric target sequence have antiparallel directionality, and said heteropolymeric probe sequence is bonded to said heteropolymeric target sequence by homologous base interaction.

9. The complex of claim 3, wherein said heteropolymeric probe sequence and said heteropolymeric target sequence have antiparallel directionality, and said heteropolymeric probe sequence is bonded to said heteropolymeric target sequence by Watson-Crick complementary base interaction.

10. The complex of claim 3, wherein:

(a) said heteropolymeric probe sequence and said heteropolymeric target sequence have antiparallel directionality;

(b) said target includes a first strand containing said heteropolymeric target sequence and a second strand containing a second heteropolymeric target sequence complementary and antiparallel to said first heteropolymeric target sequence; and

(c) said heteropolymeric probe sequence is bonded to said heteropolymeric target sequence by homologous base interaction and said heteropolymeric probe sequence is bonded to said second heteropolymeric target sequence by Watson-Crick complementary base interaction.

11. The complex of claim 1, wherein said probe and said target are double-stranded and said complex is a quadruplex.

12. The complex of claim 11, wherein said complex is free of PNA.

13. The complex of claim 11, wherein said heteropolymeric probe sequence and said heteropolymeric target sequence have parallel directionality, and said heteropolymeric probe sequence is bonded to said heteropolymeric target sequence by homologous base interaction.

14. The complex of claim 11, wherein said heteropolymeric probe sequence and said heteropolymeric target sequence have parallel directionality, and said heteropolymeric probe sequence is bonded to said heteropolymeric target sequence by Watson-Crick complementary base interaction.

15. The complex of claim 11, wherein:

(a) said heteropolymeric probe sequence and said heteropolymeric target sequence have parallel directionality;

(b) said probe includes a first probe strand containing said heteropolymeric probe sequence and a second probe strand containing a second heteropolymeric probe sequence complementary and antiparallel to said first heteropolymeric probe sequence;

(c) said target includes a first target strand containing said heteropolymeric target sequence and a second target strand containing a second heteropolymeric target sequence complementary and antiparallel to said first heteropolymeric target sequence; and

(d) said heteropolymeric probe sequence is bonded to said heteropolymeric target sequence by homologous base interaction and said heteropolymeric probe sequence is optionally bonded to said second heteropolymeric target sequence by Watson-Crick complementary base interaction.

16. The complex of claim 11, wherein:

(a) said heteropolymeric probe sequence and said heteropolymeric target sequence have parallel directionality;

(b) said probe includes a first probe strand containing said heteropolymeric probe sequence and a second probe strand containing a second heteropolymeric probe sequence complementary and antiparallel to said first heteropolymeric probe sequence;

(c) said target includes a first target strand containing said heteropolymeric target sequence and a second target strand containing a second heteropolymeric target sequence complementary and antiparallel to said first heteropolymeric target sequence; and

(d) said heteropolymeric probe sequence is bonded to said heteropolymeric target sequence by Watson-Crick complementary base interaction and said heteropolymeric probe sequence is optionally bonded to said second heteropolymeric target sequence by homologous base interaction.

17. The complex of claim 11, wherein:

(a) said heteropolymeric probe sequence and said heteropolymeric target sequence have antiparallel directionality;

(b) said probe includes a first probe strand containing said heteropolymeric probe sequence and a second probe strand containing a second heteropolymeric probe sequence complementary and antiparallel to said first heteropolymeric probe sequence;

(c) said target includes a first target strand containing said heteropolymeric target sequence and a second target strand containing a second heteropolymeric target sequence complementary and antiparallel to said first heteropolymeric target sequence; and

(d) said heteropolymeric probe sequence is bonded to said heteropolymeric target sequence by homologous base interaction and said heteropolymeric probe sequence is optionally bonded to said second heteropolymeric target sequence by Watson-Crick complementary base interaction.

18. The complex of claim 11, wherein:

(a) said heteropolymeric probe sequence and said heteropolymeric target sequence have antiparallel directionality;

(b) said probe includes a first probe strand containing said heteropolymeric probe sequence and a second probe strand containing a second heteropolymeric probe sequence complementary and antiparallel to said first heteropolymeric probe sequence;

(c) said target includes a first target strand containing said heteropolymeric target sequence and a second target strand containing a second heteropolymeric target sequence complementary and antiparallel to said first heteropolymeric target sequence; and

(d) said heteropolymeric probe sequence is bonded to said heteropolymeric target sequence by Watson-Crick complementary base interaction and said heteropolymeric probe sequence is optionally bonded to said second heteropolymeric target sequence by homologous base interaction.

19. The complex of claim 1, wherein said probe occupies a minor groove of said target.

20. The complex of claim 1, wherein said probe occupies a major groove of said target.

21. The complex of claim 3, wherein bases of said probe interact with bases of both strands of said target simultaneously.

22. The complex of claim 11, wherein bases of said probe interact with bases of both strands of said target simultaneously.

23. The complex of claim 1, wherein a backbone of the probe comprises deoxyribose phosphate.

24. The complex of claim 1, wherein a backbone of the probe is uncharged, partially negatively charged, or positively charged.

25. The complex of claim 1, wherein at least one base in said probe is a synthetic analogue of a naturally-occurring base having specific binding affinities based on Watson-Crick complementarity or homologous binding.

26. The complex of claim 1, wherein:

(a) the probe further contains a loop and a stem adjacent to the loop;

(b) the heteropolymeric probe sequence is located partially or entirely within the stem; and

(c) each of the loop and the stem comprises nucleic acids or nucleic acid analogues.

27. A method for making a complex of claim 1, said method comprising:

providing a sample comprising the target containing the heteropolymeric target sequence of nucleic acids or nucleic acid analogues;

providing the probe, wherein;

(a) the probe further contains a loop and a stem adjacent to the loop;

(b) the heteropolymeric probe sequence is located within at least one of the loop and the stem; and

(c) each of the loop and the stem comprises nucleic acids or nucleic acid analogues; and

forming a reaction mixture comprising the probe and the target under conditions that the probe and the target hybridize to form the complex.

28. The method of claim 27, further comprising detecting a signal correlated with a binding affinity between the probe and the target such that the target is detected.

29. The method of claim 28, wherein a match or a mismatch between bases of the heteropolymeric probe sequence and bases of the heteropolymeric target sequence is detected.

30. The method of claim 27, wherein the probe and the target hybridize to form a duplex.

31. The method of claim 27, wherein one of the heteropolymeric probe sequence and the heteropolymeric target sequence is single-stranded, the other of the heteropolymeric probe sequence and the heteropolymeric target sequence is double-stranded, and the probe and the target hybridize to form a triplex.

32. The method of claim 27, wherein the heteropolymeric probe sequence and the heteropolymeric target sequence are double-stranded and the probe and the target hybridize to form a quadruplex.

33. The method of claim 27, wherein:

(a) homologous binding conditions are provided and an incompletely homologous probe does not bind to the target or binds to the target with reduced efficiency;

or (b) Watson-Crick complementary binding conditions are provided and an incompletely complementary probe does not bind to the target or binds to the target with reduced efficiency.

34. The method of claim 27, wherein the heteropolymeric probe sequence is located in the stem.

35. The method of claim 27, wherein the heteropolymeric probe sequence is located in the loop.

36. The method of claim 35, wherein:

(a) the stem comprises at least two strands of nucleic acids or nucleic acid analogues, (b) the at least two strands are adapted to be bound to each other in the absence of the heteropolymeric target sequence binding to the heteropolymeric probe sequence, and (c) the at least two strands are adapted to separate from each other when the heteropolymeric target sequence binds to the heteropolymeric probe sequence.

37. The method of claim 36, wherein:

(a) one of the at least two strands comprises a donor;

(b) an opposing one of the at least two strands comprises an acceptor; and

(c) the method further comprises detecting a change in a signal when the heteropolymeric target sequence binds to the heteropolymeric probe sequence and the at least two strands separate from each other.

38. The method of claim 37, wherein:

(a) the donor emits the signal and the acceptor quenches the signal;

or (b) the donor transfers energy to the acceptor and the acceptor emits the signal.

39. The method of claim 38, wherein the energy transferred is fluorescent resonance energy.

40. The method of claim 38, wherein the signal is chemiluminescent.

41. The complex of claim 1, wherein:

(a) the probe is a heteropolymeric multistranded parallel complex;

(b) bases of the probe are homologously bound in a first plane;

(c) the target is a heteropolymeric multistranded parallel complex;

and (d) bases of the target are homologously bound in a second plane.

42. The complex of claim 1, wherein:

(a) the probe is a heteropolymeric multistranded parallel complex;

(b) bases of the probe are bound pursuant to a Watson-Crick motif in a first plane;

(c) the target is a heteropolymeric multistranded parallel complex; and

(d) bases of the target are bound pursuant to a Watson-Crick motif in a second plane.

43. An isolated or synthetic probe comprising:

a loop containing nucleic acids or nucleic acid analogues;

a stem adjacent to the loop, and containing at least two strands of nucleic acids or nucleic acid analogues; and

a heteropolymeric probe sequence located entirely or partially within the loop, and containing a double or triple-stranded sequence of nucleic acids or nucleic acid analogues,

wherein the at least two strands of the stem are adapted to be bound to each other in the absence of the heteropolymeric target sequence binding to the heteropolymeric probe sequence, and the at least two strands are adapted to separate from each other and open the loop when a heteropolymeric target sequence binds to the heteropolymeric probe sequence.