Hybridization compositions and methods
The invention provides methods and compositions for hybridizing at least one molecule to a target. The invention may, for example, eliminate the use of, or reduce the dependence on formamide in hybridization. Compositions for use in the invention include an aqueous composition comprising at least one nucleic acid sequence and at least one polar aprotic solvent in an amount effective to denature double-stranded nucleotide sequences.
1. A hybridization composition comprising a mixture of at least one nucleic acid sequence, at least one polar aprotic solvent in an amount effective to denature double-stranded nucleotide sequences, an accelerating agent, and at least one additional component selected from the group consisting of: buffering agents, salts, chelating agents, detergents, and blocking agents,
wherein less than 10% of the hybridization composition is formamide,
wherein the polar aprotic solvent is not dimethyl sulfoxide (DMSO) and has lactone, sulfone, sulfite, and/or carbonate functional groups,
wherein the at least one nucleic acid sequence is a FISH or CISH probe;
wherein the at least one polar aprotic solvent comprises a polar aprotic solvent selected from the group consisting of:
and
wherein the accelerating agent is a polymer.
2. The hybridization composition according to claim 1 , wherein the accelerating agent is present at a concentration from 1%-80%.
3. The hybridization composition according to claim 1 , wherein the accelerating agent is selected from the group consisting of PVP, heparin, dextran sulfate, and combinations thereof.
4. The hybridization composition according to claim 1 , wherein the accelerating agent is dextran sulfate.
5. The hybridization composition according to claim 4 , wherein the composition comprising up to 10% dextran sulfate.
6. The hybridization composition according to claim 4 , wherein the composition comprises up to 20% dextran sulfate.
7. The hybridization composition according to claim 1 , wherein the concentration of polar aprotic solvent is 5% to 10% (v/v).
8. The hybridization composition according to claim 1 , wherein the concentration of polar aprotic solvent is 10% to 20% (v/v).
9. The hybridization composition according to claim 1 , wherein the concentration of polar aprotic solvent is 20% to 30% (v/v).
10. The hybridization composition according to claim 1 , with the proviso that the composition does not contain formamide.
11. The hybridization composition according to claim 1 , wherein the composition contains less than 2% formamide.
12. The hybridization composition according to claim 1 wherein the composition contains less than 1% formamide.
13. The hybridization composition according to claim 1 , wherein the polar aprotic solvent is selected from the group consisting of: 2,3-butylene carbonate, γ-butyrolactone, caprolactone (epsilon), chloro maleic anhydride, chloroethylene carbonate, chloronitromethane, citraconic anhydride, crotonlactone, 5-cyano-2-thiouracil, dimethyl sulfate, dimethyl sulfone, diphenyl sulfone, ethanesulfonylchloride, ethylene carbonate, ethylene glycol sulfate, glycol sulfite, methyl alpha bromo tetronate, methyl phenyl sulfone, methyl sulfolane, methyl-4-toluenesulfonate, N-phenyl sydnone, phthalic anhydride, 1,3-propane sultone, β-propiolactone, propylene carbonate, saccharin, sulfanilamide, sulfolane, trimethylene sulfide-dioxide, and trimethylene sulfite.
14. The hybridization composition according to claim 1 , wherein the hybridization composition comprises at least one buffering agent and at least one salt.
15. The hybridization composition according to claim 14 , wherein the salt is NaCl and/or the buffering agent is phosphate buffer.
16. The hybridization composition according to claim 15 , wherein the accelerating agent is dextran sulfate present at a concentration of 10% to 40%, and wherein the NaCl is present at a concentration of 0 mM to 1200 mM, and/or the phosphate buffer is present at a concentration of 0 mM to 50 mM.
17. The hybridization composition according to claim 15 , wherein the dextran sulfate is present at a concentration of 10% to 30%, the NaCl is present at a concentration of 300 mM to 600 mM, and/or the phosphate buffer is present at a concentration of 5 mM to 20 mM.
18. The hybridization composition according to claim 1 , wherein the at least one nucleic acid sequence is a FISH DNA probe.
19. The hybridization composition according to claim 1 , wherein the at least one nucleic acid sequence comprises a detectable label.