The point of care testing of herpesvirus infection by LAMP method
| Project/Area Number |
16K08947
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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 |
Laboratory medicine
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| Research Institution | Fujita Health University |
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Principal Investigator |
Ihira Masaru 藤田医科大学, 保健学研究科, 教授 (10290165)
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| Co-Investigator(Kenkyū-buntansha) |
榎本 喜彦 藤田医科大学, 医学研究科, 研究員 (00387713)
杉山 博子 藤田医科大学, 医学研究科, 研究員 (10387714)
東本 祐紀 藤田医科大学, 医療科学部, 助教 (20569701)
吉川 哲史 藤田医科大学, 医学部, 教授 (80288472)
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| Project Period (FY) |
2016-04-01 – 2019-03-31
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| Project Status |
Completed (Fiscal Year 2018)
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| Budget Amount *help |
¥4,680,000 (Direct Cost: ¥3,600,000、Indirect Cost: ¥1,080,000)
Fiscal Year 2018: ¥1,820,000 (Direct Cost: ¥1,400,000、Indirect Cost: ¥420,000)
Fiscal Year 2017: ¥1,560,000 (Direct Cost: ¥1,200,000、Indirect Cost: ¥360,000)
Fiscal Year 2016: ¥1,300,000 (Direct Cost: ¥1,000,000、Indirect Cost: ¥300,000)
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| Keywords | LAMP / PSR / Eprobe / HHV-6 / VZV / Point of Care testing / RPA / HDA / HSV / point of care test |
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| Outline of Final Research Achievements |
The LAMP method which need six primers has problem to difficult select of primer for sensitive detection methods. In order to solve this problem, we proposed combining recombinase polymerase amplification (RPA) and the LAMP. Lower detection limits of this method(R-LAMP) to detect HHV-6 DNA were 10 copies/ reaction．Next, polymerase spiral reaction (PSR) have been developed to detect VZV DNA. The VZV PSR method could detect 10 copies / reaction of VZV DNA and more sensitive than LAMP. However, turbidity of no template control (NTC) increases by primer dimer. To discriminate between increase of turbidity by primer dimer and DNA amplification, Eprobe was used in PSR. The detection limit of PSR with Eprobe was 10 copies / reaction. However, an increase in the fluorescence intensity of NTC was confirmed by amplification of VZV DNA using PSR with Eprobe. Melting temperature analysis indicated that there was a clear difference in peak fluorescence temperature between NTC and PSR products.
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| Academic Significance and Societal Importance of the Research Achievements |
サンプルから病原体遺伝子を検出することにより迅速診断することは重要である。近年、本邦で発明された迅速に病原遺伝子増幅するLAMP方法がよく使われるようになった。LAMP法は優れた遺伝子増幅法であるが、感度に優れた検出法を構築するのは経験と時間を要する。これらの問題に対し本研究は、感度に優れたLAMP法を簡易に構築することを目的に他の遺伝子増幅法（RPA法）との組み合わせ、または近年報告されたLAMPとは異なる等温増幅方法（PSR法）による検出法を検討した。これらによって簡易に高感度検出法が構築出来れば、遺伝子診断によるPOCTとして臨床でより広く利用できるの可能性がある。
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Report
(4 results)
Research Products
(22 results)
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[Journal Article] Monitoring shedding of five genotypes of RotaTeq vaccine viruses by genotype-specific real-time RT-PCR assays.2018
Author(s)
RT-PCR assays. Higashimoto Y, Ihira M, Miyazaki Y, Kuboshiki A, Yoshinaga S, Hiramatsu H, Suzuki R, Miyata M, Miura H, Komoto S, Yukitake J, Taniguchi K, Kawamura Y, Yoshikawa T.
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Journal Title
J Clin Microbiol
Volume: 印刷中
Issue: 6
Pages: 589-596
DOI
Related Report
Peer Reviewed / Open Access
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