Triple-helix forming oligonucleotides for targeted mutagenesis
AssigneeYale University
InventorPeter M. Glazer
Life of this patent
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Abstract
A high affinity, triplex-forming oligonucleotide and methods for use thereof wherein an oligonucleotide is used to form a triple-stranded nucleic acid molecule with a specific DNA segment of a target DNA molecule. Upon formation of the triplex, the binding of the oligonucleotide stimulates mutagenesis within or adjacent to the target sequence using cellular DNA synthesis or repair mechanisms. The mutation activates, inactivates or alters the activity and function of the target molecule.
Claims11
A mutagenic composition comprising a carrier suitable for administration to cells and
- an effective amount of a high affinity oligonucleotide for targeted mutagenesis of a double-stranded nucleic acid molecule, the single-stranded oligonucleotide having a sequence that forms a triple-stranded nucleic acid molecule with a target region of the double-stranded nucleic acid molecule adjacent to a site where a mutation in the target region induced by triple helix formation will activate, inactivate or alter the activity or function of the double-stranded nucleic acid molecule or the protein it encodes, wherein the oligonucleotide has a K.sub.d of 2.times.10.sup.-8 or less for the target region, in combination with a recombination fragment.
A method of producing a high affinity, triple-helix forming oligonucleotide composition comprising the step of synthesizing an oligonucleotide substantially complementary, based on the third strand binding code, to a target region of a double-stranded nucleic acid molecule adjacent to a site where a mutation in the target region will activate, inactivate or alter the activity or function of the double-stranded nucleic acid molecule or the protein it encodes, wherein the disassociation constant for the oligonucleotide and the target region is less than or equal to 2.times.10.sup.-8, and adding to the triple-helix forming oligonucleotide a recombination fragment.
Dependencies are inferred from the claim text.