Hybridization-triggered fluorescent detection of nucleic acids
AssigneeEpoch Biosciences, Inc.
InventorsRobert O. Dempcy, Irina Aleksandrovna Afonina, Nicolaas M. J. Vermeulen
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Abstract
Compositions and methods for fluorescent detection of nucleic acids are provided. The compositions can be detected by fluorescence when hybridized to a nucleic acid containing a target sequence, but are non-fluorescent in the non-hybridized state. Alternatively, the fluorescence properties of the compositions change in a detectable manner upon hybridization to a nucleic acid containing a target sequence. Methods for synthesis and methods of use of the compositions are also provided.
Claims45
A composition comprising:
- (a) an oligonucleotide (ODN), wherein the oligonucleotide comprises a plurality of nucleotides, a 3′ end and a 5′ end;
- (b) a minor groove binder (MGB), wherein the minor groove binder is a radical of a molecule having a molecular weight of approximately 150 to approximately 5000 Daltons which molecule binds in a non-intercalating manner into the minor groove of non-single-stranded DNA, RNA or hybrids thereof with an association constant greater than approximately 103M−1;
- and
- (c) a latent fluorophore (LF);
- wherein the latent fluorophore is a radical of a molecule having a molecular weight of approximately 150 to approximately 5000 Daltons which binds in an intercalating manner into non-single-stranded DNA, RNA or hybrids thereof, or alternatively lies in the minor groove, or in another manner is oriented to the DNA molecule by the minor groove binder moiety so that it becomes fluorescent or its fluorescence properties are changed in a detectable way;
- wherein the composition is a covalently bound oligonucleotide, minor groove binder, latent fluorophore combination having a first linking group and a second linking group.
The method according to claim 18, wherein the composition of step (a) is the ODN-MGB-LF of claim 36.
The method according to claim 23, wherein the composition of step (a) is the ODN-MGB-LF of claim 36.
The method according to claim 26, wherein the composition of step (a) is the ODN-MGB-LF of claim 36.
The method according to claim 28, wherein the composition of step (d) is the ODN-MGB-LF of claim 36.
A composition comprising:
- wherein X has a backbone chain length between 0 and 48 atoms long, wherein the backbone contains atoms selected from C, N, O and S, and wherein the backbone optionally contains one or more of —NH—, —O—, —NH—C(═O)—, —C═(O)—NH—, —NH—C(═O)—NH—, —NH—C(═S)—NH—, —SO2—, —SO2—NH—, —S—or —S—S— groups;
- and wherein Y is a leaving group.
A composition comprising:
- wherein K is a cleavable linker;
- m is between 1 and 99;
- and
- n is between 1 and 47.
Dependencies are inferred from the claim text.