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US 4,820,630 AGranted
Assay for nucleic acid sequences, particularly genetic lesions, using interactive labels
AssigneeDigene Diagnostics, Incorporated
InventorFloyd Taub
Life of this patent
Estimating the term…
Abstract
Lesions in genetic sequences, for example, the sickle cell anemia mutation in the beta globin gene, are detected by means of interactive labels in a nucleic acid hybridization assay. A signal is generated only if the labels are in physical proximity which is made dependent on whether a predetermined normal or abnormal sequence is present by exposure to appropriate conditions such as restriction enzyme digestion or stringent hybridization.
Claims25
A method of detecting the presence or absence of a site for a restriction endonuclease enzyme, which comprises:
- (a) providing a sample nucleic acid to be detected for a site for a restriction endonuclease enzyme;
- (b) exposing said sample nucleic acid to a restriction endonuclease enzyme;
- (c) stably hybridizing said sample nucleic acid with
- (i) a first labeled nucleic acid probe capable of stably hybridizing to a region 5' to said site to be detected, and
- (ii) a second labeled nucleic acid probe capable of stably hybridizing to a region 3' to said site to be detected,
- the labels in said first and second nucleic acids probes together constituting a signal-generating system wherein said labels are in sufficient physical proximity so as to be capable of interacting with each other to yield a detectable signal when said site is absent but are in insufficient physical proximity so as to be less capable of yielding a detectable signal when said site is present;
- and
- (d) detecting said signal generating system, to thereby detect the presence or absence of said site.
5 more elements
A method of detecting the presence or absence of a site for a restriction endonuclease enzyme, which comprises:
- (a) providing a sample nucleic acid to be detected for a site for a restriction endonuclease enzyme;
- (b) stably hybridizing said sample nucleic acid with a doubly labeled nucleic acid probe capable of stably hydribizing to a region spanning said site when said site is absent, and carrying a first label at a position 5' to said site and a second label at a position 3' to said site;
- said first and second labels together constituting a signal generating system wherein said labels are in sufficient physical proximity so as to be capable of interacting with each other to yield a detectable signal when said site is absent but are in insufficient physical proximity so as to be less capable of yielding a detectable signal when said site is present;
- (c) exposing said sample nucleic acid to a restriction endonuclease enzyme;
- and
- (d) detecting said signal generating system, to thereby detect the presence or absence of said site.
3 more elements
A method of detecting the presence or absence of a region of biological significance in a nucleic acid sequence, which comprises:
- (a) providing a sample nucleic acid sequence to be detected for the presence or absence of a region of biological significance in said sequence,
- (b) incubating said sample nucleic acid with
- (i) a first labelled nucleic acid probe capable of stably hydridizing to said nucleic acid sequence when said region is absent, and
- (ii) a second labelled nucleic acid probe capable of stably hydribizing to a region proximate thereto;
- wherein both said first and second labels are enzymes, and said labels interact by diffusion of a chemical species from one to another;
- the enzymes in said first and second nucleic acid probes together constituting a signal generating system, wherein said enzymes are in sufficient physical proximity so as to be capble of interacting with each other to yield a detectable signal when said region of biological signficance is absent, but are in insufficient physical proximity so as to be less capable of yielding a detectable signal when said region of biological significance is present;
- and
- (c) detecting said signal generating system to thereby detect the presence or absence of said region to biological significance.
5 more elements
A method of detecting the presence or absence of a region of biological signifance in a nucleic acid sequence, which comprises:
- (a) providing a sample nucleic acid sequence to be detected for the presence or absence of a region of biological significance in said sequence,
- (b) incubating said sample nucleic acid with
- (i) a first labelled nucleic acid probe capability of stably hybridizing to said nucleic acid sequence when said region is present, and
- (ii) a second labelled nucleic acid probe capable of stably hydridizing to a region proximate thereto;
- wherein both said first and second labels are enzymes and said enzymes interact by diffusion of a chemical species from one to another;
- the enzymes in said first and second nucleic acid probes together constituting a signal generating system, wherein said enzymes are in sufficient physical proximity so as to be capable of interacting with each other to yield a detectable signal when said region of biological significance is present, but are in insufficient proximity so as to be less capable of yielding a detectable signal when said region of biological significance is absent;
- and
- (c) detecting said signal generating system to thereby detect the presence or absence of said region of biological significance.
5 more elements
A method of detecting the presence or absence of a site for a restriction endonuclease enzyme which comprises,
- (a) providing a sample nucleic acid containing a site for a restriction endonuclease enzyme in a genetic sequence;
- (b) labeling said sequence with a first label;
- (c) exposing said sample nucleic acid to a restriction endonuclease enzyme;
- (d) hybridizing said sample nucleic acid with a nucleic acid probe capable of stably hybridizing to the region spanning said site when said site is absent, said probe labeled with a second label,
- said first and second labels together constituting a signal generating signal wherein said labels are in sufficient physical proximity so as to be capable of interacting with each other to yield a detectable signal when said site is absent, but are in insufficient physical proximity so as to be less capable of yielding a signal when said site is present;
- and
- (e) detecting said signal generating system to thereby detect the presence or absence of said site.
4 more elements
The method of claims 1, 4, 7, 79, or 80 wherein said sample nucleic acid is first attached to a support means.
Dependencies are inferred from the claim text.