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AssigneesThe University of Tokyo, Nikon Corporation
InventorsTaro Ueno, Takashi Funatsu, Takanori Ichiki, Hirofumi Shiono
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A method for detecting a target nucleic acid, comprising: (a) contacting a nucleic acid sample comprising a target nucleic acid, comprising a first portion and a second portion, with: (i) a detection probe, wherein the detection probe is labeled with a labeling substance and comprises a nucleic acid sequence that forms a stem-loop structure and having a 5′ protruding end or a 3′ protruding end that is capable of hybridizing to the second portion, and (ii) a capture probe comprising a nucleic acid sequence capable of hybridizing to the first portion, wherein the capture probe is immobilized to a substrate, under conditions to form a target nucleic acid-detection probe-capture probe complex by hybridizing the second portion to the detection probe and hybridizing the first portion to the capture probe; (b) ligating a first end of the detection probe with an end of the target nucleic acid and ligating a second end of the detection probe with an end of the capture probe; and (c) detecting the labeling substance of the nucleic acid-detection probe-capture probe complex formed on the substrate.
A method for detecting a target nucleic acid, comprising:
Dependencies are inferred from the claim text.
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Involving nucleic acids: nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes
Hybridisation assays: enzymatic or biochemical coupling of nucleic acids to a solid phase
Enzymatic or biochemical coupling of nucleic acids to a solid phase: using probe arrays or probe chips
Oligonucleotides characterized by their use: mirna, sirna or ncrna
About 7 years 2 months of term left · ends 16 Dec 2033 (estimated)
The instant application contains a Sequence Listing which has been submitted electronically in ASCII format and is hereby incorporated by reference in its entirety. Said ASCII copy, created on Oct. 30, 2014, is named 107929-0105_SL.txt and is 8,686 bytes in size.
The present invention relates to a nucleic acid detection method, detection probe, detection probe set, and nucleic acid quantification method.
Conventionally, DNA microarray targeting mRNA is widely used as a means of measuring the gene expression levels in cells. In the 21st century, it is reported that miRNA, Filch is a short-chain and a non-coding RNA, controls gene expression in vivo, and relationships between abnormal expression of miRNA and a variety of diseases such as the cancer have been elucidated. Based on these findings, development race of the DNA microarray targeting miRNA takes place.
FIG. 1 is a schematic view of one embodiment of a method for quantifying nucleic acids in this embodiment.
FIG. 2 is a schematic view of one embodiment of the detection probe in the embodiment.
FIG. 3 is a quantitative result of miRNA in Example 1.
<<Method for Detecting Nucleic Acids>>
The nucleic acid to be detected (target nucleic acid) is not particularly limited, short chain RNA such as miRNA or short chain mRNA generated by stopping transcription on the way is preferable and miRNA which exists in various types in vivo, and is involved in the regulation of gene expression is more preferable.
As an example, a method for quantifying nucleic acids of the present embodiment comprises, (a) a step of contacting a solution comprising: a nucleic acid sample containing miRNA comprising a first portion and a second portion, and a detection probe labeled with a labeling substance containing a sequence which forms a stem-loop structure and is capable of hybridizing to the second portion, and having 5′ protruding end or 3′ protruding end, with a substrate fixed with a capture probe containing a sequence capable of hybridizing to the first portion, (b) a step of forming the miRNA—the detection probe—the capture probe complex on the substrate by hybridizing the second portion to the detection probe and hybridizing the first portion to the capture probe, (c) a step of ligating an end of the detection probe and ends of the miRNA and the capture probe, (d) a step of quantitatively detecting the labeling substance of the miRNA—the detection probe—the capture probe complex formed on the substrate, and quantifying miRNA of the nucleic acid sample from a detection result.