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US 8,137,937 B2Granted
Method for bisulfite treatment
AssigneeRoche Molecular Systems, Inc.
InventorChristine Markert-Hahn
Life of this patent
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Abstract
The present application is directed to a method for performing a bisulfite reaction to determine methylation positions in a nucleic acid via treatment of the solid phase-bound nucleic acid with bisulfite, desulfonation and elution of the nucleic acid from the solid phase.
Claims16
A method for the conversion of a cytosine base in a nucleic acid to an uracil base comprising the steps of
- a1) binding the nucleic acid to a solid phase capable of binding nucleic acids and incubating the solid phase bound nucleic acid in the presence of sulfite ions thereby deaminating the cytosine base in the nucleic acid, or
- a2) incubating the nucleic acid in the presence of sulfite ions thereby deaminating the cytosine base in the nucleic acid and binding the nucleic acid to a solid phase,
- b) optionally washing the solid phase bound nucleic acid,
- c) contacting the solid phase bound nucleic acid with a solution with an alkaline pH thereby desulfonating the base deaminated in step a1) or a2) in the nucleic acid and converting it to an uracil base and simultaneously eluting the nucleic acid.
1 more element
A method for the amplification of a nucleic acid comprising the steps of
- a1) binding the nucleic acid to a solid phase capable of binding nucleic acids and incubating the solid phase bound nucleic acid in the presence of sulfite ions thereby deaminating the cytosine base in the nucleic acid, or
- a2) incubating the nucleic acid in the presence of sulfite ions thereby deaminating the cytosine base in the nucleic acid and binding the nucleic acid to a solid phase,
- b) optionally washing the solid phase bound nucleic acid,
- c) contacting the solid phase bound nucleic acid with a solution with an alkaline pH thereby
- desulfonating the base deaminated in step a1) or a2) in the nucleic acid and converting it to an uracil base and
- simultaneously eluting the nucleic acid,
- d) amplifying the eluted nucleic acid by PCR or related method.
4 more elements
A kit for performing a bisulfite reaction comprising
- a bisulfite salt or a solution comprising bisulfite ions,
- a solid phase capable of binding nucleic acids, and
- a solution comprising 10 to 100 mM NaOH or KOH.
Dependencies are inferred from the claim text.