Multichromophore fluorescent probes using DNA intercalation complexes
AssigneeThe Regents of the University of California
InventorsAlexander N. Glazer, Richard A. Mathies, Konan Peck
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Abstract
Novel fluorescent labeling techniques and fluorescent labels are provided, employing high affinity non-covalently binding and intercalating fluorescent dyes and dsDNA. The dyes find application to provide highly sensitive labeling of nucleic acids in electrophoretic gels and as pre-prepared labels for binding to a wide variety of specific binding pair members. The DNA-dye fluorescer complex can be used for labels in diagnostic assays, detection of specific nucleic acid sequences, and the like.
Claims19
A fluorescent label comprising (a) a nucleic acid of at least about 20 nucleotides and (b) non-covalently bound and intercalated in said nucleic acid at least one fluorescent molecule characterized by having at least two positive charges, having at least one fluorescent monomeric unit and having a binding affinity to double stranded DNA of at least about 5×106 M−1 and up to about 1 fluorescent unit per 4 nucleotides in a nucleic acid strand,
- wherein said fluorescent unit is characterized by having at least one aromatic ring per monomeric unit, and
- wherein said nucleic acid comprises a member of a specific binding pair capable of binding to a complementary member of said specific binding pair or a functionality for covalent bonding to another molecule other than a naturally occurring functionality of a nucleotide.
A fluorescent label comprising:
- (a) double stranded DNA,
- (b) a fluorescent aromatic dimer having two monomeric fluorescent units joined by a chain of at least about 10 Å non-covalently bound and intercalated in said double stranded DNA at a ratio of at least about 1:5 base pairs, and
- (c) covalently joined to said double stranded DNA a ligand member of a specific binding pair.
A composition comprising:
- (a) a double stranded DNA of at least about 20 base pairs and
- (b) intercalated and non-covalently bound in said double stranded DNA at least one fluorescent molecule of at least two positive charges, having at least one fluorescent monomeric unit and having a binding affinity to double stranded DNA of at least about 5×106 M−1 and up to about 1 fluorescent unit per 4 nucleotides in a nucleic acid strand, wherein said fluorescent unit is characterized by having at least one aromatic ring per monomeric unit, wherein said double stranded DNA is covalently linked to a polypeptide or saccharide.
Dependencies are inferred from the claim text.