| Project/Area Number |
18K09003
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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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| Review Section |
Basic Section 56010:Neurosurgery-related
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| Research Institution | Saitama Medical University |
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Principal Investigator |
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| Co-Investigator(Kenkyū-buntansha) |
西川 亮 埼玉医科大学, 医学部, 教授 (90237678)
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| Project Period (FY) |
2018-04-01 – 2021-03-31
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| Project Status |
Completed (Fiscal Year 2020)
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| Budget Amount *help |
¥2,730,000 (Direct Cost: ¥2,100,000、Indirect Cost: ¥630,000)
Fiscal Year 2020: ¥650,000 (Direct Cost: ¥500,000、Indirect Cost: ¥150,000)
Fiscal Year 2019: ¥910,000 (Direct Cost: ¥700,000、Indirect Cost: ¥210,000)
Fiscal Year 2018: ¥1,170,000 (Direct Cost: ¥900,000、Indirect Cost: ¥270,000)
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| Keywords | Glioma / TERT / ddPCR / glioma / droplet digital PCR / digital PCR / 遺伝子変異 / プロモーター / HRM / mutation |
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| Outline of Final Research Achievements |
It is important to accurately detect TERT promoter mutations in glioma. Sanger DNA sequencing is the currently standard method for analyzing TERT mutations. In this report, we described a novel droplet digital PCR (ddPCR) assay to evaluate TERT hot spot mutations in fresh frozen and formalin-fixed paraffin-embedded (FFPE) specimens of glioma and verified the difference in results from the Sanger DNA sequencing results. We obtained the mutant allele fraction for TERT mutations of in a single ddPCR run in all cases, including the microdissected FFPE sections. On the contrary, up to twice the DNA sequences were required from fresh frozen tissue to obtain the results, consistent with ddPCR assay. When FFPE specimens were used, more time was required to evaluate TERT mutations through DNA sequencing. DdPCR is an effective and sensitive assay compared to the conventional standard Sanger DNA sequencing.
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| Academic Significance and Societal Importance of the Research Achievements |
ddPCR法はサンプル内にPCR増幅される遺伝子があれば必ず増幅検出されるため、0.01％の変異DNAでも検出が可能な高感度な遺伝子変異検出法であり再現性も高い。特に変異のホットスポットが判明しているTERT遺伝子変異の解析に対しては極めて有用な方法である。
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