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METHOD FOR DETECTING HUMAN GENOMIC DNA NEW

外国特許コード F180009500
整理番号 (S2017-0155-N0)
掲載日 2018年11月1日
出願国 世界知的所有権機関(WIPO)
国際出願番号 2017JP042943
国際公開番号 WO 2018101375
国際出願日 平成29年11月30日(2017.11.30)
国際公開日 平成30年6月7日(2018.6.7)
優先権データ
  • 特願2016-233119 (2016.11.30) JP
発明の名称 (英語) METHOD FOR DETECTING HUMAN GENOMIC DNA NEW
発明の概要(英語) Provided is a method for detecting and/or assaying human genomic DNA in a test sample at high sensitivity. One "human Alu model sequence" representing the consensus sequence of 46 human Alu subfamilies is identified, and PCR primer-probe candidates for Alu detection are selected using this human Alu model sequence. Next, a primer-probe set that is an oligonucleotide sequence which hybridizes specifically with the human Alu sequence and that comprises an oligonucleotide sequence capable of minimizing dimer formation is selected using proprietary criteria. The human genomic DNA in the test sample can be detected and/or assayed at high sensitivity by conducting quantitative real-time PCR using the PCR primer-probe for Alu detection obtained in this manner.
従来技術、競合技術の概要(英語) BACKGROUND ART
Alu sequences, retrotransposon SINE(short interspersed element) in a kind of, including a human primate genome specific iterative sequence is present.Alu sequences, referred to as left monomer and light monomer 2 from one sequence 7SL RNA, sandwiched by them is made from the A-rich sequence.Alu sequences in the genome of a primate is a specific sequence, constitute a part of the 7SL RNA derived sequences are also present in the genome of the rodent is known.In addition, a recent study, Alu sequences in the human genome consists of a sequence of all is not the same, sub-families 46 is clear (non-patent document 1).
Alu sequences, that there be at least 100 million copies in the human genome, 10% or more of the human genome is known.A total length of approximately 30 billion nucleotides of the human genome from each pair, according to simple calculation, the average for 1 copies of DNA3000 nucleotide pairs in the human genome Alu sequence exists according to the present embodiment.Is converted into weight, of one times the weight of the whole human genome in from 3pg, 0.003fg of human genomic DNA sequences 1 included in the average is considered to be a copy of the Alu.In this way, the sequence of human genomic DNA to a very small Alu always present, further, a primate from the specific nucleotide sequence, the detection of human cells, in an ideal target for the determination of the thought.
Alu and quantitative real-time PCR method to target (hereinafter, 'Alu-qPCR' are sometimes referred to) is used to detect the human genome, is a method for the quantification, it has been reported a plurality of (for example, Patent Document 1, Non-Patent Document 2 or the like).In addition, as a kit for Alu-qPCR, Innoquant (registered trademark) (InnoGenomics Technologies Corporation) or, (registered trademark) Human DNA Quantification Kit(ZYMO RESEARCH Corporation) Femto already commercially available.Further, using Alu-qPCR, a mouse xenograft and engraftment of human stem cells, and the user can confirm the degree of growth (non-patent document 3), DNA concentration was measured in the blood of cancer diagnosis (Non-Patent Document 4) can be, the element used in the field of forensic human genomic DNA in the sample can be detected to have been reported (Non-Patent Document 5).
However, have been reported so far either Alu-qPCR not have sufficient sensitivity.That is, in theory, real-time PCR DNA from the human genome of approximately 0.1fg by Alu can be detected which should, in one or more known Alu-qPCR pg of human genomic DNA is the number required (non-patent document 5) and, compared to the theoretical limit of detection is not only a very low sensitivity.Also, in known Alu-qPCR primers, non-human species of the same portion of the genome and the nucleotide sequence, genomic DNA derived from other species of the human genome and DNA may be mixed with the case of using a sample, specifically Alu trace was impossible to detect.From the above, has a higher sensitivity and specificity of Alu-qPCR development has been demanded.
  • 出願人(英語)
  • ※2012年7月以前掲載分については米国以外のすべての指定国
  • AKITA UNIVERSITY
  • 発明者(英語)
  • AKASHI HIDEO
  • FUNAKOSHI KODAI
国際特許分類(IPC)
指定国 National States: AE AG AL AM AO AT AU AZ BA BB BG BH BN BR BW BY BZ CA CH CL CN CO CR CU CZ DE DJ DK DM DO DZ EC EE EG ES FI GB GD GE GH GM GT HN HR HU ID IL IN IR IS JO JP KE KG KH KN KP KR KW KZ LA LC LK LR LS LU LY MA MD ME MG MK MN MW MX MY MZ NA NG NI NO NZ OM PA PE PG PH PL PT QA RO RS RU RW SA SC SD SE SG SK SL SM ST SV SY TH TJ TM TN TR TT TZ UA UG US UZ VC VN ZA ZM ZW
ARIPO: BW GH GM KE LR LS MW MZ NA RW SD SL SZ TZ UG ZM ZW
EAPO: AM AZ BY KG KZ RU TJ TM
EPO: AL AT BE BG CH CY CZ DE DK EE ES FI FR GB GR HR HU IE IS IT LT LU LV MC MK MT NL NO PL PT RO RS SE SI SK SM TR
OAPI: BF BJ CF CG CI CM GA GN GQ GW KM ML MR NE SN ST TD TG
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