Nucleic acid hybridization essay method
First Claim
1. A nucleic acid hybridization assay method comprising:
- contacting a nucleic acid hybridization assay device with a liquid sample including a target nucleic acid, wherein the nucleic acid hybridization assay device comprises a cleavable signal element comprising a capture probe of a single strand having a complementary sequence to the target nucleic acid; and
a single stranded restriction probe not being complementary to the target nucleic acid, wherein one end of the single stranded restriction probe is ligated to the capture probe; and
a solid support substrate attached to the other end of the single stranded restriction probe;
hybridizing the cleavable signal element to the target nucleic acid present in the liquid sample;
making the single stranded restriction probe into a double stranded restriction probe by DNA extension in the presence of a DNA polymerization solution using the target nucleic acid hybridized to the capture probe as a primer;
cleaving the double stranded restriction probe by a restriction enzyme or the double stranded cleavable signal element by a cleavage enzyme, wherein the double stranded restriction probe is cleaved by the restriction enzyme specifically responsive to a double strand with a particular sequence and the cleavage enzyme specifically responsive to double stranded nucleic acids;
washing the nucleic acid hybridization assay device to remove the cleavable signal element cleaved by the restriction enzyme or the cleavage enzyme; and
detecting whether the uncleaved signal element or the cleaved signal element exists on the solid support substrate.
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Abstract
A cleavable signal element applicable to quantitative and qualitative assay devices, using a cleavable technique specifically responsive to a complementary double strand or single strand of nucleic acids, and a nucleic acid hybridization assay method and device using the cleavable signal element are provided. Using the cleavable technique responsive to the complementary double strand or single strand of nucleic acids, detection sensitivity to a target nucleic acid can be increased, and diagnosis and detection reliability can be improved twice through in-situ determinations. Through simultaneous single nucleotide polymorphism (SNP) detection and expression profile determination, more accurate diagnosis for many diseases can be achieved. The assay device can be easily modified to be suitable for detection with general laser-based detection systems such as CD-ROM readers. Information read from the assay device is digitized as software and transmitted to and received by doctors and patients through a computer network or wirelessly, which enables construction of remote diagnosis systems.
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Citations
10 Claims
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1. A nucleic acid hybridization assay method comprising:
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contacting a nucleic acid hybridization assay device with a liquid sample including a target nucleic acid, wherein the nucleic acid hybridization assay device comprises a cleavable signal element comprising a capture probe of a single strand having a complementary sequence to the target nucleic acid; and
a single stranded restriction probe not being complementary to the target nucleic acid, wherein one end of the single stranded restriction probe is ligated to the capture probe; and
a solid support substrate attached to the other end of the single stranded restriction probe;hybridizing the cleavable signal element to the target nucleic acid present in the liquid sample; making the single stranded restriction probe into a double stranded restriction probe by DNA extension in the presence of a DNA polymerization solution using the target nucleic acid hybridized to the capture probe as a primer; cleaving the double stranded restriction probe by a restriction enzyme or the double stranded cleavable signal element by a cleavage enzyme, wherein the double stranded restriction probe is cleaved by the restriction enzyme specifically responsive to a double strand with a particular sequence and the cleavage enzyme specifically responsive to double stranded nucleic acids; washing the nucleic acid hybridization assay device to remove the cleavable signal element cleaved by the restriction enzyme or the cleavage enzyme; and detecting whether the uncleaved signal element or the cleaved signal element exists on the solid support substrate. - View Dependent Claims (3, 4, 5, 6, 7, 8, 9, 10)
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2. A nucleic acid hybridization assay method comprising:
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contacting a nucleic acid hybridization assay device with a liquid sample including a target nucleic acid, wherein the nucleic acid hybridization assay device comprises a cleavable signal element comprising a capture probe of a single strand having a complementary sequence to the target nucleic acid; and
a single stranded restriction probe not being complementary to the target nucleic acid, wherein one end of the single stranded restriction probe is ligated to the capture probe; and
a solid support substrate attached to the other end of the single stranded restriction probe;hybridizing the cleavable signal element to the target nucleic acid present in the liquid sample; making the single stranded restriction probe remain as a single strand after DNA extension except when the single stranded restriction probe is changed into a double stranded restriction probe by the DNA extension, where the DNA extension is done in the presence of a DNA polymerization solution using the target nucleic acid hybridized to the cleavable signal element as a primer; cleaving the single stranded restriction probe by a cleavage enzyme after the DNA extension, wherein the cleavage enzyme specifically responsive to the single stranded nucleic acids; washing the nucleic acid hybridization assay device to remove the cleavable signal element cleaved by the cleavage enzyme; and detecting whether the uncleaved signal element or the cleaved signal element exists on the solid support substrate.
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Specification