Title: Detection of nucleic acids by multiple sequential invasive cleavages

Abstract

The present invention relates to means for the detection and characterization of nucleic acid sequences, as well as variations in nucleic acid sequences. The present invention also relates to methods for forming a nucleic acid cleavage structure on a target sequence and cleaving the nucleic acid cleavage structure in a site-specific manner. The structure-specific nuclease activity of a variety of enzymes is used to cleave the target-dependent cleavage structure, thereby indicating the presence of specific nucleic acid sequences or specific variations thereof. The present invention further relates to methods and devices for the separation of nucleic acid molecules based on charge. The present invention also provides methods for the detection of non-target cleavage products via the formation of a complete and activated protein binding region. The invention further provides sensitive and specific methods for the detection of human cytomegalovirus nucleic acid in a sample.

Inventors:

Hall, Jeff G ^[1]; Lyamichev, Victor I ^[1]; Mast, Andrea L ^[1]; Brow, Mary Ann D ^[1]

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1. Madison, WI

Issue Date:

1999-01-01

Research Org.:

Third Wave Technologies, Inc., Madison, WI (United States)

OSTI Identifier:

872708

Patent Number(s):

5994069

Assignee:

Third Wave Technologies, Inc. (Madison, WI)

Patent Classifications (CPCs):

C - CHEMISTRY C12 - BIOCHEMISTRY C12Q - MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS

C - CHEMISTRY C12 - BIOCHEMISTRY C12N - MICROORGANISMS OR ENZYMES

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C - CHEMISTRY C12 - BIOCHEMISTRY C12Q - MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS

C12Q1/6823 - Release of bound markers
C12Q2525/186 - incorporating a non-extendable or blocking moiety
C12Q2525/131 - incorporating a restriction site
C12Q2561/109 - Invader technology
C12Q1/6827 - for detection of mutation or polymorphism
C12Q2521/301 - Endonuclease

C - CHEMISTRY C12 - BIOCHEMISTRY C12N - MICROORGANISMS OR ENZYMES

C12N9/1252 - {DNA-directed DNA polymerase

C - CHEMISTRY C12 - BIOCHEMISTRY C12Q - MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS

C12Q1/683 - involving restriction enzymes, e.g. restriction fragment length polymorphism [RFLP]
C12Q2525/143 - incorporating a promoter sequence
C12Q1/6816 - characterised by the detection means
C12Q1/70 - involving virus or bacteriophage
C12Q2525/155 - incorporating/generating a new priming site
C12Q2537/149 - Sequential reactions
C12Q1/701 - {Specific hybridization probes}

C - CHEMISTRY C12 - BIOCHEMISTRY C12N - MICROORGANISMS OR ENZYMES

C12N9/22 - Ribonucleases {RNAses, DNAses

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DOE Contract Number:  

FG02-94ER81891

Resource Type:

Patent

Country of Publication:

United States

Language:

English

Subject:

detection; nucleic; acids; multiple; sequential; invasive; cleavages; relates; means; characterization; acid; sequences; variations; methods; forming; cleavage; structure; target; sequence; cleaving; site-specific; manner; structure-specific; nuclease; activity; variety; enzymes; cleave; target-dependent; indicating; presence; specific; devices; separation; molecules; based; charge; provides; non-target; products; via; formation; complete; activated; protein; binding; region; sensitive; human; cytomegalovirus; sample; acid molecule; provides methods; cleavage structure; acid molecules; acid sequence; nucleic acid; acid sequences; nucleic acids; target sequence; acid cleavage; specific nucleic; structure-specific nuclease; molecules based; specific variations; site-specific manner; nuclease activity; target-dependent cleavage; specific manner; /435/