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METHOD FOR PRODUCING GENE KNOCK-IN CELLS

外国特許コード F180009356
整理番号 (S2016-0981-N0)
掲載日 2018年4月19日
出願国 世界知的所有権機関(WIPO)
国際出願番号 2017JP027838
国際公開番号 WO 2018030208
国際出願日 平成29年8月1日(2017.8.1)
国際公開日 平成30年2月15日(2018.2.15)
優先権データ
  • 特願2016-157484 (2016.8.10) JP
発明の名称 (英語) METHOD FOR PRODUCING GENE KNOCK-IN CELLS
発明の概要(英語) The present inventor attempted to produce a knock-in animal with the use of a single-stranded DNA as a donor by utilizing a genome editing system comprising a nuclease in the form of a protein and a DNA-targeting RNA. As a result, it was found that cells wherein the donor DNA was knocked-in could be thus obtained at an extremely high efficiency.
特許請求の範囲(英語) [claim1]
1. A region of the desired target DNA DNA inserted in the method for forming a cell, a protein having an activity Nuclease (a), (b) a target DNA nucleotide sequence of a region (a) the nucleotide sequence and complementary to interact with a protein containing the base sequence of DNA IgGκ RNA, and comprising the desired DNA (c) donor-stranded DNA method comprising a step of introducing into the cell.
[claim2]
2. A target DNA desired DNA inserted into the area of the cell in which the method for forming a, (a) a protein Cas9, crRNA tracrRNA fragment (b) a combination of fragments, and (c) a single-chain comprising a desired DNA donor DNA method comprising a step of introducing into a cell.
[claim3]
3. (c) A single-stranded donor DNA base sequence having 600 or more bases of the, method according to claim 1 or 2.
[claim4]
4. (c) Is a single-stranded donor DNA, or (ii) the next (i) is prepared in a method, the method of any one of claims 1-3. (i) A pair of different amount of PCR primers by amplifying DNA, and the double-stranded DNA contained in the amplification product specifically decomposition method comprising a step of (ii) a pair of primers, on the other hand to the primer PCR primer phosphorylations by amplifying the DNA using, and the amplification product is phosphorylated at specific chain decomposition method comprising a step of
[claim5]
5. The cell is an oocyte, method according to any one of claims 1-4.
[claim6]
6. A fertilized oocytes, method according to claim 5.
[claim7]
7. Inserted into the area of the desired target DNA DNA containing the cell of the method for forming a non-human individuals, any one of claims 1-6 (a) the step of performing a method, and the step (b) (a) a non-human individual cells obtained by the steps of preparing a from, the method comprising.
[claim8]
8. A non-human mammal is a rodent, method according to claim 7.
[claim9]
9. For use in the method of any one of claims 1-8 kit, (c) from the following (a) at least one selected from the group consisting of a kit comprising a molecule 1. (a) Target DNA nuclease (b) a protein having an activity that is complementary to a nucleotide sequence of a region (a) of the nucleotide sequence and nucleotide sequence of DNA interact with a target RNA (c) including the desired DNA single-stranded DNA to the donor
  • 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
  • TOKYO MEDICAL AND DENTAL UNIVERSITY
  • 発明者(英語)
  • AIDA TOMOMI
  • TANAKA KOHICHI
国際特許分類(IPC)
指定国 (WO201830208)
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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