| Project/Area Number |
26670165
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| Research Category |
Grant-in-Aid for Challenging Exploratory Research
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| Allocation Type | Multi-year Fund |
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| Research Field |
Pathological medical chemistry
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| Research Institution | Institute of Physical and Chemical Research |
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Principal Investigator |
Takumi Toru 国立研究開発法人理化学研究所, 脳科学総合研究センター, シニアチームリーダー (00222092)
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| Project Period (FY) |
2014-04-01 – 2016-03-31
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| Project Status |
Completed (Fiscal Year 2015)
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| Budget Amount *help |
¥3,770,000 (Direct Cost: ¥2,900,000、Indirect Cost: ¥870,000)
Fiscal Year 2015: ¥1,950,000 (Direct Cost: ¥1,500,000、Indirect Cost: ¥450,000)
Fiscal Year 2014: ¥1,820,000 (Direct Cost: ¥1,400,000、Indirect Cost: ¥420,000)
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| Keywords | コピー数多型 / 染色体工学 / ゲノム編集技術 / 自閉症 / ES細胞 / ゲノム編集 / CRISPR / TALEN |
|---|
| Outline of Final Research Achievements |
In the CRISPR-Cas9 system, a single guide RNA (sgRNA) directs the endonuclease Cas9 to a targeted DNA sequence for site-specific manipulation. The sgRNA has also been shown to play a role in activating the endonuclease activity of Cas9. This dual function of the sgRNA likely underlies observations that different sgRNAs have varying on-target activities. We evaluated the cleavage activities of 218 sgRNAs using in vitro Surveyor assays. We found that nucleotides at both PAM-distal and PAM-proximal region of the sgRNA are significantly correlated with on-target efficiency. Furthermore, we demonstrated that the genomic contexts of the targeted DNA, the GC percentage, and the secondary structure of sgRNA are critical factors contributing to cleavage efficiency.
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