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Title:
FLUORESCENT QUENCHING DETECTION REAGENTS AND METHODS
Document Type and Number:
WIPO Patent Application WO2001042505
Kind Code:
A3
Abstract:
Oligonucleotide-fluorophore-quencher conjugates wherein the fluorophore moiety has emission wavelengths in the range of about (300) to about (800) nm, and or where the quencher includes a substituted 4-(phenyldiazenyl)phenylamine structure provide improved signal to noise ratios and other advantageous characteristics in hybridization and related assays. The oligonucleotide-fluorophore-quencher conjugates can be synthesized by utilizing novel phosphoramidite reagents that incorporate the quencher moiety based on the substituted 4-(phenyldiazenyl)phenylamine structure, and or novel phosphoramidite reagents that incorporate a fluorophore moiety based on the substituted coumarin, substituted 7-hydroxy-3H-phenoxazin-3-one, or substituted 5,10-dihydro-10-[phenyl]pyrido[2,3-d;6,5-d']dipyrimidine-2,4,6,8-(1H, 3H, 7H, 9H, 10H)-tetrone structure. Oligonucleotide-fluorophore-quencher-minor groove binder conjugates including a pyrrolo[4,5-e]indolin-7-yl}carbonyl)pyrrolo[4,5-e]indolin-7-yl]carbonyl}pyrrolo[4,5-e]indoline-7-carboxylate (DPI3) moiety as the minor groove binder and the substituted 4-(phenyldiazenyl)phenylamine moiety as the quencher, were synthesized and have substantially improved hybridization and signal to noise ratio properties.

Inventors:
REED MICHAEL W
LUKHTANOV EUGENY ALEXANDER
GALL ALEXANDER A
DEMPCY ROBERT O
Application Number:
US0033333W
Publication Date:
January 24, 2002
Filing Date:
December 08, 2000
Export Citation:
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Assignee:
EPOCH BIOSCIENCES, INC.
International Classes:
C12N15/09; C07C245/08; C07D519/00; C07F9/24; C07F9/572; C07F9/655; C07F9/6561; C07H21/00; C07H21/04; C09B29/08; C12Q1/68; C07B61/00; (IPC1-7): C07H21/00; C07C245/08; C07F9/24; C12Q1/68
Domestic Patent References:
WO1997039008A11997-10-23
Foreign References:
US5304645A1994-04-19
US5952202A1999-09-14
US5801155A1998-09-01
Other References:
ASANUMA H ET AL: "Photo-Responsive Oligonucleotides Carrying Azobenzene in the Side-Chains", TETRAHEDRON LETTERS,NL,ELSEVIER SCIENCE PUBLISHERS, AMSTERDAM, vol. 39, no. 49, 3 December 1998 (1998-12-03), pages 9015 - 9018, XP004140991, ISSN: 0040-4039
"'98 Catalogue", 1998, AMERSHAM PHARMACIA BIOTECH, 'S-HERTOGENBOSCH, NETHERLANDS, XP002172887
LEE L G ET AL: "ALLELIC DISCRIMINATION BZ NICK-TRANSLATION PCR WITH FLUOROGENIC PROBES", NUCLEIC ACIDS RESEARCH,GB,OXFORD UNIVERSITY PRESS, SURREY, vol. 21, no. 16, 11 August 1993 (1993-08-11), pages 3761 - 3766, XP000470188, ISSN: 0305-1048
NAZARENKO ET AL: "A CLOSED TUBE FORMAT FOR AMPLIFICATION AND DETECTION OF DNA BASED ENERGY TRANSFER", NUCLEIC ACIDS RESEARCH,GB,OXFORD UNIVERSITY PRESS, SURREY, vol. 25, no. 12, 1997, pages 2516 - 2521, XP002094959, ISSN: 0305-1048
SOKOL D L ET AL: "Real time detection of DNA.RNA hybridization in living cells", PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF USA,US,NATIONAL ACADEMY OF SCIENCE. WASHINGTON, vol. 95, no. 20, September 1998 (1998-09-01), pages 11538 - 11543, XP002123471, ISSN: 0027-8424
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