Screening sequence selectivity of oligonucleotide-binding molecules using nanoparticle based colorimetric assay

1. A method of assessing selectivity of a destabilizing oligonucleotide-binding molecule comprising:

a) heating a first mixture comprising (1) a first complex comprising a first functionalized nanoparticle and a second functionalized nanoparticle, (2) a first hairpin oligonucleotide, and (3) the destabilizing oligonucleotide-binding molecule to determine a first melting temperature of the first complex in the first mixture,

b) comparing the first melting temperature of the first complex in the first mixture to a second melting temperature of the first complex in a second mixture comprising (1) the first complex, (2) a second different hairpin oligonucleotide; and (3) the destabilizing oligonucleotide-binding molecule,

wherein (i) a higher first melting temperature than the second melting temperature indicates stronger selectivity of the destabilizing oligonucleotide-binding molecule compound for the first hairpin oligonucleotide than for the second hairpin oligonucleotide, (ii) a lower first melting temperature than second melting temperature indicates a stronger selectivity of the destabilizing oligonucleotide-binding molecule for the second hairpin oligonucleotide than for the first hairpin oligonucleotide, and (iii) a first melting temperature essentially equal to the second melting temperature indicates essentially equal selectivity of the destabilizing oligonucleotide-binding molecule for the second hairpin oligonucleotide and the first hairpin oligonucleotide; and

the first functionalized nanoparticle comprises a first oligonucleotide on at least a portion of the first nanoparticle surface and the second functionalized nanoparticle comprises a second oligonucleotide on at least a portion of the second nanoparticle surface, and the first oligonucleotide is sufficiently complementary to the second oligonucleotide to hybridize.

2. A method of assessing selectivity of a stabilizing oligonucleotide-binding molecule comprising:

a) heating a first mixture comprising (1) a first complex comprising a first functionalized nanoparticle and a second functionalized nanoparticle, (2) a first hairpin oligonucleotide, and (3) the stabilizing oligonucleotide-binding molecule to determine a first melting temperature of the first complex in the first mixture,

b) comparing the first melting temperature of the first complex in the first mixture to a second melting temperature of the first complex in a second mixture comprising (1) the first complex, (2) a second different hairpin oligonucleotide; and (3) the stabilizing oligonucleotide -binding molecule,

wherein (i) a higher first melting temperature than the second melting temperature indicates stronger selectivity of the stabilizing oligonucleotide-binding molecule compound for the second hairpin oligonucleotide than for the first hairpin oligonucleotide, (ii) a lower first melting temperature than second melting temperature indicates a stronger selectivity of the stabilizing oligonucleotide-binding molecule for the first hairpin oligonucleotide than for the second hairpin oligonucleotide, and (iii) a first melting temperature essentially equal to the second melting temperature indicates essentially equal selectivity of the stabilizing oligonucleotide-binding molecule for the second hairpin oligonucleotide and the first hairpin oligonucleotide; and

the first functionalized nanoparticle comprises a first oligonucleotide on at least a portion of the first nanoparticle surface and the second functionalized nanoparticle comprises a second oligonucleotide on at least a portion of the second nanoparticle surface, and the first oligonucleotide is sufficiently complementary to the second oligonucleotide to hybridize.

3. The method of claim 1 , wherein the first oligonucleotide or second oligonucleotide is DNA.

4. The method of claim 1 , wherein the first oligonucleotide and second oligonucleotide are DNA.

5. The method of claim 1 , wherein the first oligonucleotide or the second oligonucleotide is a modified oligonucleotide.

6. The method of claim 1 , wherein the first oligonucleotide and the second oligonucleotide are modified oligonucleotides.

7. The method of claim 1 , wherein the first hairpin oligonucleotide or the second hairpin oligonucleotide is DNA.

8. The method of claim 1 , wherein the first hairpin oligonucleotide and the second hairpin oligonucleotide are DNA.

9. The method of claim 1 , wherein the first hairpin oligonucleotide or the second hairpin oligonucleotide is a modified oligonucleotide.

10. The method of claim 1 , wherein the first hairpin oligonucleotide and the second hairpin oligonucleotide are modified oligonucleotides.

11. The method of claim 1 , wherein the first nanoparticle and the second nanoparticle are gold nanoparticle.

12. The method of claim 11 , wherein the first melting temperature is determined by detecting a blue-to-red color change associated with de-aggregation of the first gold nanoparticle and the second gold nanoparticle.

13. The method of claim 12 , wherein the color change is detected without instrumentation.

14. The method of claim 11 , wherein the first nanoparticle, second nanoparticle, or both have a diameter of about 13 nm to about 250 nm.

15. The method of claim 1 , which is performed in a high throughput format.