Non-Fluorescent Quencher Compounds and Biomolecular Assays
AssigneeLife Technologies Corporation
InventorsGregory J. Ewing, Deborah Kay Fieri, Khairuzzaman Bashar Mullah, Ronald J. Graham
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Abstract
Bis-diazo, triaryl and aryldiazo-N-arylphenazonium quencher moieties, substituted with electron-withdrawing and electron-donating substituents which induce polarity in the delocalized aryl/diazo ring systems, are useful as labels when attached to biomolecules such as polynucleotides, nucleosides, nucleotides and polypeptides. The quencher moieties are non-fluorescent and accept energy transfer from fluorescent reporter labels by any energy-transfer mechanism, such as FRET. Fluorescence quencher compositions are useful in preparing quencher labeled biomolecules for various molecular biology assays based on fluorescence detection.
Claims54
A compound comprising:
- (i) a quencher moiety comprising structure (Q):
- wherein:
- at least one aryl carbon of (Q) is substituted with an electron-withdrawing group;
- and
- at least one aryl carbon of (Q) is substituted with an electron-donating group;
- and
- (ii) an optionally protected reactive linking group linked to an aryl carbon of the quencher moiety, either directly or by way of an optional linker moiety.
A composition useful for synthesizing a quencher-labeled polynucleotide, comprising:
- (i) a quencher moiety comprising structure (Q):
- wherein:
- at least one aryl carbon of (Q) is substituted with an electron-withdrawing group;
- and
- at least one aryl carbon of (Q) is substituted with an electron-donating group;
- (ii) a hydroxyl protected with an acid-labile protecting group;
- and
- (iii) a solid support, wherein the protected hydroxyl group and the quencher moiety are linked to the solid support via an optionally cleavable linker.
A quencher-labeled conjugate, comprising a quencher moiety and a conjugated substance, wherein the quencher moiety comprises structure (Q):
- wherein:
- at least one aryl carbon is substituted with an electron-withdrawing group;
- and
- at least one aryl carbon is substituted with an electron-donating group,
- and further wherein the quencher moiety is covalently linked to the conjugated substance at an aryl carbon, optionally by way of a linker.
Dependencies are inferred from the claim text.