| Project/Area Number |
15K06902
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| Research Category |
Grant-in-Aid for Scientific Research (C)
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| Allocation Type | Multi-year Fund |
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| Section | 一般 |
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| Research Field |
Genome biology
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| Research Institution | Kindai University |
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Principal Investigator |
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| Co-Investigator(Renkei-kenkyūsha) |
KOHCHI Takayuki 京都大学, 大学院生命科学研究科, 教授 (40202056)
ISHIZAKI Kimitsune 神戸大学, 大学院理学研究科, 准教授 (00452293)
TAKIKAWA Yoshihiro 近畿大学, 先端技術総合研究所, 准教授 (60446010)
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| Project Period (FY) |
2015-04-01 – 2018-03-31
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| Project Status |
Completed (Fiscal Year 2017)
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| Budget Amount *help |
¥4,940,000 (Direct Cost: ¥3,800,000、Indirect Cost: ¥1,140,000)
Fiscal Year 2017: ¥1,560,000 (Direct Cost: ¥1,200,000、Indirect Cost: ¥360,000)
Fiscal Year 2016: ¥1,820,000 (Direct Cost: ¥1,400,000、Indirect Cost: ¥420,000)
Fiscal Year 2015: ¥1,560,000 (Direct Cost: ¥1,200,000、Indirect Cost: ¥360,000)
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| Keywords | RNA-protein complex / CRISPR/Cas9 / microinjection / ゼニゴケ / マイクロインジェクション / ゲノム編集 / RNA-タンパク質複合体 |
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| Outline of Final Research Achievements |
DNA-free genome editing allows target-specific manipulation of a given genome. CRISPR RNA and Cas9 protein can be directly introduced into target cells and lead to genome-editing. We have successfully performed DNA-free genome editing mediated by microinjection of CRISPR/Cas9 complex in the liverwort Marchantia polymorpha. NOPPERABO1 (NOP1), of which loss-of-function mutation causes impaired air-chamber formation and thus can be found readily, was selected as a target gene. Custom gRNA and commercially available Cas9 protein were first allowed to form RNA-protein complex and then injected into single-cell sporelings by a laser thermal microinjector. One of the thalli grown from microinjected sporelings formed a sector that showed the nop1 mutant phenotype, and DNA isolated from the sector showed 5-bp deletion in the target coding sequence. This is the first demonstration of DNA-free genome-editing by microinjection in plants.
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