Claims
- 1. A method for identifying at least one homozygous genetic factor associated with a first phenotype that differs from a second phenotype, wherein an association between the at least one homozygous genetic factor and the first phenotype is unknown, comprising:
providing at least one strand of first-phenotype knockout DNA substantially devoid of polymorphisms and at least one strand of second-phenotype knockout DNA substantially devoid of polymorphisms; performing a denaturing/reannealing cycle on a pool comprising the at least one strand of first-phenotype knockout DNA and the at least one strand of second-phenotype knockout DNA; and capturing at least one mismatch in the pool following the denaturing/reannealing cycle, the captured at least one mismatch representing the at least one homozygous genetic factor associated with the first phenotype.
- 2. The method according to claim 1, wherein providing at least one strand of first-phenotype knockout DNA comprises:
providing at least one strand of first-phenotype DNA; digesting the at least one strand of first-phenotype DNA with four cutter restriction enzymes to produce a plurality of first-phenotype DNA fragments; performing at least one cycle of denaturing/reannealing on the plurality of first-phenotype DNA fragments; treating a plurality of reannealed first-phenotype DNA fragments with at least one endonuclease such that one or more mismatched DNA regions in the plurality of reannealed first-phenotype DNA fragments are removed without modifying DNA sequences adjacent the one or more mismatched DNA regions to produce the at least one strand of first-phenotype knockout DNA substantially devoid of polymorphisms.
- 3. The method according to claim 1, wherein providing the at least one strand of second-phenotype knockout DNA comprises:
providing at least one strand of second-phenotype DNA; digesting the at least one strand of second-phenotype DNA with four cutter restriction enzymes to produce a plurality of second-phenotype DNA fragments; performing at least one cycle of denaturing/reannealing on the plurality of second-phenotype DNA fragments; treating a plurality of reannealed second-phenotype DNA fragments with at least one endonuclease such that one or more mismatched DNA regions in the plurality of reannealed second-phenotype DNA fragments are removed without modifying DNA sequences adjacent the one or more mismatched DNA regions to produce the at least one strand of second-phenotype knockout DNA substantially devoid of polymorphisms.
- 4. The method according to claim 1, further comprising:
performing a subsequent denaturing/reannealing cycle on a subsequent pool comprising the at least one strand of uncaptured first-phenotype knockout DNA and the at least one strand of uncaptured second-phenotype knockout DNA; and capturing at least one mismatch in the subsequent pool following the subsequent denaturing/reannealing cycle, the at least one mismatch representing the at least one homozygous genetic factor associated with the first phenotype.
- 5. The method according to claim 2, wherein the plurality of first-phenotype DNA fragments average less than approximately 500 base pairs long.
- 6. The method according to claim 5, wherein the plurality of first-phenotype DNA fragments average approximately 300-400 base pairs long.
- 7. The method according to claim 3, wherein the plurality of second-phenotype DNA fragments average less than approximately 500 base pairs long.
- 8. The method according to claim 7, wherein the plurality of second-phenotype DNA fragments average approximately 300-400 base pairs long.
- 9. The method according to claim 1, wherein the first phenotype is at least one of a disease predisposition, a disease susceptibility, a drug reaction predisposition and a behavior predisposition.
- 10. The method according to claim 1, wherein the at least one homozygous genetic factor comprises a polymorphism.
- 11. The method according to claim 1, wherein the at least one homozygous genetic factor comprises a single nucleotide polymorphism.
- 12. The method according to claim 1, wherein the at least one homozygous genetic factor comprises a plurality of polymorphisms.
- 13. The method according to claim 1, wherein the at least one homozygous genetic factor comprises a plurality of single nucleotide polymorphisms.
- 14. The method according to claim 1, wherein capturing at least one mismatch comprises exposing at least one mismatch to a DNA repair protein.
- 15. The method according to claim 14, wherein the DNA repair protein is of a type that binds to mismatched DNA.
- 16. The method according to claim 14, wherein the DNA repair protein is a MutS protein.
- 17. The method according to claim 1, further comprising differentially labeling the at least one strand of first-phenotype knockout DNA and the at least one strand of second-phenotype knockout DNA.
- 18. The method according to claim 1, further comprising characterizing the captured at least one mismatch.
- 19. The method according to claim 18, wherein characterizing the captured at least one mismatch comprises sequencing the at least one mismatch.
- 20. The method according to claim 19, further comprising performing bioinformatic analysis on the mismatch.
- 21. A method for diagnosing a disease associated with at least one homozygous genetic factor, comprising identifying the at least one homozygous genetic factor associated with the disease by the method according to claim 1.
- 22. A method for predicting a drug response associated with at least one homozygous genetic factor, comprising identifying the at least one homozygous genetic factor associated with the drug response by the method according to claim 1.
- 23. A method for predicting a disease predisposition associated with at least one homozygous genetic factor, comprising identifying the at least one homozygous genetic factor associated with the disease predisposition by the method according to claim 1.
- 24. A method for predicting a disease susceptibility associated with at least one homozygous genetic factor, comprising identifying the at least one homozygous genetic factor associated with the disease susceptibility by the method according to claim 1.
- 25. A method for identifying at least one heterozygous genetic factor associated with a first phenotype that differs from a second phenotype, wherein an association between the at least one heterozygous genetic factor and the first phenotype is unknown, comprising:
providing at least one strand of first-phenotype capture DNA enriched for polymorphisms of interest and at least one strand of second-phenotype knockout DNA substantially devoid of polymorphisms; performing a denaturing/reannealing cycle on a pool comprising at least one strand of first-phenotype capture DNA and at least one strand of second-phenotype knockout DNA; and capturing at least one mismatch in the pool following the denaturing/reannealing cycle, the at least one mismatch representing the at least one heterozygous genetic factor associated with the first phenotype.
- 26. The method according to claim 25, wherein providing at least one strand of first-phenotype capture DNA comprises:
providing at least one strand of first-phenotype DNA; digesting the at least one strand of first-phenotype DNA with four cutter restriction enzymes to produce a plurality of first-phenotype DNA fragments; and capturing a plurality of first-phenotype mismatched DNA fragments in at least one cycle such that one or more mismatched DNA regions in the plurality of first-phenotype DNA fragments is retained to produce at least one strand of first-phenotype capture DNA enriched with polymorphisms of interest.
- 27. The method according to claim 25, wherein providing at least one strand of second-phenotype knockout DNA comprises:
providing at least one strand of second-phenotype DNA; digesting at least one strand of second-phenotype DNA with four cutter restriction enzymes to produce a plurality of second-phenotype DNA fragments; performing at least one cycle of denaturing/reannealing on the plurality of second-phenotype DNA fragments; treating a plurality of reannealed second-phenotype DNA fragments with at least one endonuclease such that one or more mismatched DNA regions in the plurality of reannealed second-phenotype DNA fragments are removed without modifying DNA sequences adjacent the one or more mismatched DNA regions to produce at least one strand of second-phenotype knockout DNA substantially devoid of polymorphisms.
- 28. The method according to claim 25, further comprising:
performing a subsequent denaturing/reannealing cycle on a subsequent pool comprising at least one strand of uncaptured first-phenotype capture DNA and at least one strand of uncaptured second-phenotype knockout DNA; and capturing at least one mismatch in the subsequent pool following the subsequent denaturing/reannealing cycle, at least one mismatch representing at least one heterzygous genetic factor associated with the first phenotype.
- 29. The method according to claim 27, wherein the plurality of second-phenotype DNA fragments average less than approximately 500 base pairs long.
- 30. The method according to claim 29, wherein the plurality of second-phenotype DNA fragments average approximately 300-400 base pairs long.
- 31. The method according to claim 25, wherein the first phenotype comprises at least one of a disease predisposition, a disease susceptibility, a drug reaction predisposition and a behavior predisposition.
- 32. The method according to claim 25, wherein at least one heterozygous genetic factor comprises a polymorphism.
- 33. The method according to claim 25, wherein at least one heterozygous genetic factor comprises a single nucleotide polymorphism.
- 34. The method according to claim 25, wherein at least one heterozygous genetic factor comprises a plurality of polymorphisms.
- 35. The method according to claim 25, wherein at least one heterozygous genetic factor comprises a plurality of single nucleotide polymorphisms.
- 36. The method according to claim 25, wherein capturing at least one mismatch comprises exposing the at least one mismatch to a DNA repair protein.
- 37. The method according to claim 36, wherein the DNA repair protein is of a type that binds to mismatched DNA.
- 38. The method according to claim 37, wherein the DNA repair protein is a MutS protein.
- 39. The method according to claim 25, further comprising differentially labeling the at least one strand of first-phenotype capture DNA and the at least one strand of second-phenotype knockout DNA.
- 40. The method according to claim 25, further comprising characterizing the captured at least one mismatch.
- 41. The method according to claim 40, wherein characterizing the captured at least one mismatch comprises sequencing the at least one mismatch.
- 42. The method according to claim 41, further comprising performing bioinformatic analysis on the captured at least one mismatch.
- 43. A method for diagnosing a disease associated with at least one heterozygous genetic factor, comprising identifying the at least one heterozygous genetic factor associated with the disease by the method according to claim 25.
- 44. A method for predicting a drug response associated with at least one heterozygous genetic factor, comprising identifying the at least one heterozygous genetic factor associated with the drug response by the method according to claim 25.
- 45. A method for predicting a disease predisposition associated with at least one heterozygous genetic factor, comprising identifying the at least one heterozygous genetic factor associated with the disease predisposition by the method according to claim 25.
- 46. A method for predicting a disease susceptibility associated with at least one heterozygous genetic factor, comprising identifying the at least one heterozygous genetic factor associated with the disease susceptibility by the method according to claim 25.
- 47. An apparatus for identifying at least one homozygous genetic factor associated with a first phenotype that differs from a second phenotype, wherein an association between the at least one homozygous genetic factor and the first phenotype is unknown, the apparatus comprising:
a knockout DNA-producing component for producing at least one strand of first-phenotype knockout DNA substantially devoid of polymorphisms and at least one strand of second-phenotype knockout DNA substantially devoid of polymorphisms; a DNA denaturing/reannealing component for performing a denaturing/reannealing cycle on a pool comprising at least one strand of first-phenotype knockout DNA and at least one strand of second-phenotype knockout DNA; and a capture component for capturing at least one mismatch in the pool following the denaturing/reannealing cycle, the captured at least one mismatch representing at least one homozygous genetic factor associated with the first phenotype.
- 48. A system for identifying at least one homozygous genetic factor associated with a first phenotype that differs from a second phenotype, wherein an association between the at least one homozygous genetic factor and the first phenotype is unknown, the system comprising:
a knockout DNA-producing component for producing at least one strand of first-phenotype knockout DNA substantially devoid of polymorphisms and at least one strand of second-phenotype knockout DNA substantially devoid of polymorphisms; a DNA denaturing/reannealing component for performing a denaturing/reannealing cycle on a pool comprising at least one strand of first-phenotype knockout DNA and at least one strand of second-phenotype knockout DNA; a capture component for capturing at least one mismatch in the pool following the denaturing/reannealing cycle, the captured at least one mismatch representing at least one homozygous genetic factor associated with the first phenotype; and a processing component in communication with the knockout DNA-producing component, the DNA denaturing/reannealing component, and the capture component, the processing component providing monitoring and control of the functions of the knockout DNA-producing component, the DNA denaturing/reannealing component, and the capture component.
- 49. An apparatus for identifying at least one heterozygous genetic factor associated with a first phenotype that differs from a second phenotype, wherein an association between the at least one heterozygous genetic factor and the first phenotype is unknown, the apparatus comprising:
a capture DNA/knockout DNA-producing component for producing at least one strand of first-phenotype capture DNA enriched for polymorphisms of interest and at least one strand of second-phenotype knockout DNA substantially devoid of polymorphisms; a DNA denaturing/reannealing component for performing a denaturing/reannealing cycle on a pool comprising at least one strand of first-phenotype capture DNA and at least one strand of second-phenotype knockout DNA; and a capture component for capturing at least one mismatch in the pool following the denaturing/reannealing cycle, the captured at least one mismatch representing at least one heterozygous genetic factor associated with the first phenotype.
- 50. A system for identifying at least one heterozygous genetic factor associated with a first phenotype that differs from a second phenotype, wherein an association between the at least one heterozygous genetic factor and the first phenotype is unknown, the system comprising:
a capture DNA/knockout DNA-producing component for producing at least one strand of first-phenotype capture DNA enriched for polymorphisms of interest and at least one strand of second-phenotype knockout DNA substantially devoid of polymorphisms; a DNA denaturing/reannealing component for performing a denaturing/reannealing cycle on a pool comprising at least one strand of first-phenotype capture DNA and at least one strand of second-phenotype knockout DNA; a capture component for capturing at least one mismatch in the pool following the denaturing/reannealing cycle, the captured at least one mismatch representing at least one heterozygous genetic factor associated with the first phenotype; and a processing component in communication with the knockout DNA-producing component, the DNA denaturing/reannealing component, and the capture component, the processing component providing monitoring and control of the functions of the knockout DNA-producing component, the DNA denaturing/reannealing component, and the capture component.
- 51. The method according to claim 1, wherein providing at least one strand of first-phenotype knockout DNA substantially devoid of polymorphisms and the at least one strand of second-phenotype DNA substantially devoid of polymorphisms comprises providing DNA fragments, genomic DNA, cDNA, cloned genes, or other genetic material.
- 52. The method according to claim 25, wherein providing at least one strand of first-phenotype capture DNA enriched for polymorphisms of interest and at least one strand of second-phenotype knockout DNA substantially devoid of polymorphisms comprises providing DNA fragments, genomic DNA, cDNA, cloned genes, or other genetic material.
Parent Case Info
[0001] This application claims the benefit of co-pending provisional application Ser. No. 60/450,606; filed Feb. 26, 2003, the entirety of which is incorporated herein by this reference.
Provisional Applications (1)
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Number |
Date |
Country |
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60450606 |
Feb 2003 |
US |