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Development of quantitative method by loop-mediated isothermal amplification (LAMP) using cycling probe.

Research Project

Project/Area Number 25460705
Research Category

Grant-in-Aid for Scientific Research (C)

Allocation TypeMulti-year Fund
Section一般
Research Field Laboratory medicine
Research InstitutionFujita Health University

Principal Investigator

IHIRA Masaru  藤田保健衛生大学, 保健学研究科, 教授 (10290165)

Co-Investigator(Kenkyū-buntansha) SUGIYAMA Hiroko  藤田保健衛生大学, 医学研究科, 研究員 (10387714)
ENOMOTO Yoshihiko  藤田保健衛生大学, 医学研究科, 研究員 (00387713)
YOSHIKAWA Tetsushi  藤田保健衛生大学, 医学部, 教授 (80288472)
Project Period (FY) 2013-04-01 – 2016-03-31
Project Status Completed (Fiscal Year 2015)
Budget Amount *help
¥5,070,000 (Direct Cost: ¥3,900,000、Indirect Cost: ¥1,170,000)
Fiscal Year 2015: ¥1,950,000 (Direct Cost: ¥1,500,000、Indirect Cost: ¥450,000)
Fiscal Year 2014: ¥1,690,000 (Direct Cost: ¥1,300,000、Indirect Cost: ¥390,000)
Fiscal Year 2013: ¥1,430,000 (Direct Cost: ¥1,100,000、Indirect Cost: ¥330,000)
KeywordsHHV-6A / HHV-6B / LAMP / cycling probe / Real-timePCR / Cycling probe / Taqman probe / HHV-6 / real-time PCR
Outline of Final Research Achievements

Aim of this study was to develop quantitative LAMP method for measurement of target DNA load by using cycling probe. We combine LAMP and cycling probe to develop new quantitative method. The cycling probe PCR had a broad, linear dynamic range of detection between 10 and 1000000 copies of viral DNA. Strong correlations were demonstrated between cycling probe PCR and Taqman real-time PCR. The optimized condition for cycling probe LAMP was confirmed using variable concentration of pH and probe. The cycling probe LAMP assay had excellent linear dynamic range between 1000 and 1000000 copies of viral DNA. However, cycling probe LAMP was less sensitive than Taqman real-time PCR. Result of detection from HHV6 DNA in serum by our developed dry reagent for LAMP were correspond with that of liquid reagent. Our cycling probe LAMP require improved sensitivity. However, combine dry reagent-LAMP and cycling probe suggest the possibility of rapid and simple DNA quantitation method.

Report

(4 results)
  • 2015 Annual Research Report   Final Research Report ( PDF )
  • 2014 Research-status Report
  • 2013 Research-status Report
  • Research Products

    (31 results)

All 2016 2015 2014 2013 Other

All Journal Article (17 results) (of which Peer Reviewed: 17 results,  Acknowledgement Compliant: 2 results,  Open Access: 7 results) Presentation (14 results) (of which Int'l Joint Research: 3 results)

  • [Journal Article] A simple cytogenetic method to detect chromosomally integrated human herpesvirus-6.2016

    • Author(s)
      Ohye T, et al.
    • Journal Title

      J Virol Methods

      Volume: 228 Pages: 74-78

    • DOI

      10.1016/j.jviromet.2015.11.001

    • Related Report
      2015 Annual Research Report
    • Peer Reviewed / Open Access
  • [Journal Article] Cycling probe-based real-time PCR for the detection of human herpesvirus 6A and B.2016

    • Author(s)
      Ihira M, Yamaki A, Kato Y, Higashimoto Y, Kawamura Y, Yoshikawa T.
    • Journal Title

      J Med Virol.

      Volume: Epub ahead of print Issue: 9 Pages: 000-000

    • DOI

      10.1002/jmv.24513

    • Related Report
      2015 Annual Research Report
    • Peer Reviewed / Acknowledgement Compliant
  • [Journal Article] Pathogenic Role of Human Herpesvirus 6B Infection in Mesial Temporal Lobe Epilepsy.2015

    • Author(s)
      Kawamura Y, Nakayama A, Kato T, Miura H, Ishihara N, Ihira M, Takahashi Y, Matsuda K, Yoshikawa T.
    • Journal Title

      Journal of Infectious disease

      Volume: 印刷中 Issue: 7 Pages: 1014-21

    • DOI

      10.1093/infdis/jiv160

    • Related Report
      2015 Annual Research Report
    • Peer Reviewed / Acknowledgement Compliant
  • [Journal Article] Pathogenesis of Severe Neutropenia in Patients With Primary Human Herpesvirus 6B Infection.2015

    • Author(s)
      Miura H, Kawamura Y, Ozeki E, Ihira M, Ohashi M, Yoshikawa T.
    • Journal Title

      Pediatr Infect Dis J.

      Volume: 34(9) Issue: 9 Pages: 1003-7

    • DOI

      10.1097/inf.0000000000000777

    • Related Report
      2015 Annual Research Report
    • Peer Reviewed
  • [Journal Article] Virological analysis of inherited chromosomally integrated human herpesvirus-6 in three hematopoietic stem cell transplant patients.2015

    • Author(s)
      Miura H, Kawamura Y, Kudo K, Ihira M, Ohye T, Kurahashi H, Kawashima N, Miyamura K, Yoshida N, Kato K, Takahashi Y, Kojima S, Yoshikawa T.
    • Journal Title

      Transpl Infect Dis.

      Volume: 17 Issue: 5 Pages: 728-731

    • DOI

      10.1111/tid.12419

    • Related Report
      2015 Annual Research Report
    • Peer Reviewed / Open Access
  • [Journal Article] Nationwide survey of rotavirus-associated encephalopathy and sudden unexpected death in Japan.2014

    • Author(s)
      Kawamura Y, Ohashi M, Ihira M, Hashimoto S, Taniguchi K, Yoshikawa T.
    • Journal Title

      Brain & Development

      Volume: 36 Pages: 601-607

    • Related Report
      2014 Research-status Report
    • Peer Reviewed
  • [Journal Article] Virus specific cell-mediated immunity may play a role in controlling reactivated human herpesvirus 6B in patients under measles induced immunosuppression.2014

    • Author(s)
      Kumagai T, Yoshikawa T, Shiraki K, Yoshida M, Nakayama T, Ihira M, Asano Y.
    • Journal Title

      Journal of medical virology

      Volume: 86 Issue: 4 Pages: 658-665

    • DOI

      10.1002/jmv.23862

    • Related Report
      2014 Research-status Report 2013 Research-status Report
    • Peer Reviewed
  • [Journal Article] Copy numbers of telomeric repeat sequences of human herpesvirus 6B in clinical isolates: possibility of mixed infections.2014

    • Author(s)
      Kato Y, Ihira M, Umeda M, Higashimoto Y, Kawamura Y, Ohashi M, Ishi J, Yoshikawa T.
    • Journal Title

      J Clin Microbiol.

      Volume: 52(2) Issue: 2 Pages: 419-424

    • DOI

      10.1128/jcm.02192-13

    • Related Report
      2014 Research-status Report 2013 Research-status Report
    • Peer Reviewed / Open Access
  • [Journal Article] Direct detection of human herpesvirus 6B by the LAMP method using newly developed dry-reagents.2014

    • Author(s)
      Yoshikawa T, Matsuo T, Kawamura Y, Ohashi M, Yonekawa T, Kanda H, Notomi T, Ihira M.
    • Journal Title

      Journal of Virological methods

      Volume: 201 Pages: 65-67

    • DOI

      10.1016/j.jviromet.2014.02.017

    • Related Report
      2014 Research-status Report
    • Peer Reviewed / Open Access
  • [Journal Article] Dual roles for the telomeric repeats in chromosomally integrated human herpesvirus-6.2014

    • Author(s)
      Ohye T, Inagaki H, Ihira M, Higashimoto Y, Kato K, Oikawa J, Yagasaki H, Niizuma T, Takahashi Y, Kojima S, Yoshikawa T, Kurahashi H
    • Journal Title

      Sci Rep

      Volume: 4 Issue: 1 Pages: 4559-4559

    • DOI

      10.1038/srep04559

    • Related Report
      2014 Research-status Report
    • Peer Reviewed / Open Access
  • [Journal Article] The kinetics of urinary shedding of BK virus in children with renal disease.2014

    • Author(s)
      Yamamoto Y, Morooka M, Ihira M, Yoshikawa T.
    • Journal Title

      Microbiology and immunology

      Volume: 59 Issue: 1 Pages: 37-42

    • DOI

      10.1111/1348-0421.12212

    • NAID

      40020351702

    • Related Report
      2014 Research-status Report
    • Peer Reviewed / Open Access
  • [Journal Article] Analysis of the shedding of three β-herpesviruses in urine and saliva of children with renal disease.2014

    • Author(s)
      Yamamoto Y, Morooka M, Hashimoto S, Ihira M, Yoshikawa T.
    • Journal Title

      J Med Virol.

      Volume: 86(3) Issue: 3 Pages: 505-11

    • DOI

      10.1002/jmv.23782

    • Related Report
      2013 Research-status Report
    • Peer Reviewed / Open Access
  • [Journal Article] Direct detection of human herpesvirus 6B by the LAMP method using newly developed dry-reagents.2014

    • Author(s)
      Yoshikawa T, Matsuo T, Kawamura Y, Ohashi M, Yonekawa T, Kanda H, Notomi T, Ihira M.
    • Journal Title

      J Virol Methods.

      Volume: 201 Pages: 65-7

    • Related Report
      2013 Research-status Report
    • Peer Reviewed
  • [Journal Article] Quantitative analysis of shedding of Epstein-Barr virus in saliva from patients with connective tissue diseases: a pilot study.2013

    • Author(s)
      Kuhara T, Watanabe D, Ishida N, Tamada Y, Matsumoto Y, Ihira M, Fukaya S, Yoshida S, Yoshikawa T, Asano Y.
    • Journal Title

      Int J Dermatol.

      Volume: 52(7) Issue: 7 Pages: 887-90

    • DOI

      10.1111/j.1365-4632.2011.04992.x

    • Related Report
      2013 Research-status Report
    • Peer Reviewed
  • [Journal Article] Cycling probe technology to quantify and discriminate between wild-type varicella-zoster virus and Oka vaccine strains.2013

    • Author(s)
      Ihira M, Higashimoto Y, Kawamura Y, Sugata K, Ohashi M, Asano Y, Yoshikawa T.
    • Journal Title

      J Virol Methods.

      Volume: 29 Issue: 3 Pages: 308-313

    • DOI

      10.1002/jmv.23788

    • Related Report
      2013 Research-status Report
    • Peer Reviewed
  • [Journal Article] Nationwide survey of rotavirus-associated encephalopathy and sudden unexpected death in Japan.2013

    • Author(s)
      Kawamura Y, Ohashi M, Ihira M, Hashimoto S, Taniguchi K, Yoshikawa T.
    • Journal Title

      Brain Dev.

      Volume: S0387-7604(13) Issue: 7 Pages: 234-9

    • DOI

      10.1016/j.braindev.2013.07.013

    • Related Report
      2013 Research-status Report
    • Peer Reviewed
  • [Journal Article] Clinical utility of loop-mediated isothermal amplification assay for the diagnosis of common alpha herpesvirus skin infections.2013

    • Author(s)
      Kobayashi T, Yagami A, Suzuki K, Ihira M, Yoshikawa T, Matsunaga K.
    • Journal Title

      J Dermatol.

      Volume: 40(12) Issue: 12 Pages: 1033-7

    • DOI

      10.1111/1346-8138.12325

    • Related Report
      2013 Research-status Report
    • Peer Reviewed
  • [Presentation] Development of HHV-6A Real-Time RT-PCR Assays for Detection ofThree Classes of HHV-6A Gene Transcripts2015

    • Author(s)
      Masaru Ihira, Hiroki Miura, Yuki Higashimoto, Fumihiko Hattori, Yoshiki Kawamura, Ken Sugata, Tetsushi Yoshikaw
    • Organizer
      9th international conference on HHV-6,7
    • Place of Presentation
      Boston MA,USA
    • Year and Date
      2015-11-09
    • Related Report
      2015 Annual Research Report
    • Int'l Joint Research
  • [Presentation] Human Herpesvirus 6B Infection in Pediatric Hematopoietic Stem Cell Transplantation.2015

    • Author(s)
      Hiroki Miura1, Yoshiki Kawamura, Fumihiko Hattori1, Kazuko Kudo, Masaru Ihira, Tetsushi Yoshikawa
    • Organizer
      9th international conference on HHV-6,7
    • Place of Presentation
      Boston MA,USA
    • Year and Date
      2015-11-09
    • Related Report
      2015 Annual Research Report
    • Int'l Joint Research
  • [Presentation] The Analysis of the Origin of Chromosomally Integrated Human Herpesvirus 62015

    • Author(s)
      Yoshiki Kawamura1, Tamae Ohye2, Hiroki Miura, Masaru Ihira, Hiroki Kurahashi, Tetsushi Yoshikawa
    • Organizer
      9th international conference on HHV-6,7
    • Place of Presentation
      Boston MA,USA
    • Year and Date
      2015-11-09
    • Related Report
      2015 Annual Research Report
    • Int'l Joint Research
  • [Presentation] Real-time RT PCR法によるHHV-6AmRNA検出法の構築2015

    • Author(s)
      井平 勝、東本 祐紀 、加藤友理、平松裕之、鈴木竜太、三浦浩樹、河村吉紀、吉川 哲史
    • Organizer
      第56回日本臨床ウイルス学会
    • Place of Presentation
      日本 岡山
    • Year and Date
      2015-06-13
    • Related Report
      2015 Annual Research Report
  • [Presentation] LAMP 法によるMycobacterium tuberculosis特異的 rans-renal DNA検出 第2報2015

    • Author(s)
      井平 勝、中島千絵、浅野喜造、鈴木定彦、吉川哲史
    • Organizer
      第7回LAMP研究会
    • Place of Presentation
      東京 蒲田 富士通ソリューションスクエア
    • Year and Date
      2015-02-22
    • Related Report
      2014 Research-status Report
  • [Presentation] Pathogenic role of humam herpesvirus 6B in mesial temporal sclerosis2014

    • Author(s)
      Yoshiki kawamura Hiriki miura Masaru Ihira Yukitishi Takahashi Kazumi Matsuda Tetsushi Yoshikawa
    • Organizer
      Annual International Herpesvirus Workshop
    • Place of Presentation
      Japan Kobe convention center
    • Year and Date
      2014-07-19 – 2014-07-24
    • Related Report
      2014 Research-status Report
  • [Presentation] Human herpes virus 6A and 6B specific real-time PCR by using cycling probe technology2014

    • Author(s)
      Masaru Ihira Yuri Kato Yuki Higashimoto Hiroyuki Hiramatsu Hiroko Sugiyama Yoshihiko Enomoto Yoshiki Kawamura Ryota Suzuki Tetsushi Yoshikawa
    • Organizer
      Annual International Herpesvirus Workshop
    • Place of Presentation
      Japan Kobe convention center
    • Year and Date
      2014-07-19 – 2014-07-24
    • Related Report
      2014 Research-status Report
  • [Presentation] Cycling probeを用いたhuman herpesvirus 6 A,Bの新規DNA定量法開発2014

    • Author(s)
      井平 勝,榎本喜彦 ,杉山博子 ,東本 祐紀 ,平松裕之, 鈴木竜太 ,河村吉紀 ,吉川 哲史
    • Organizer
      第54回日本臨床ウイルス学会
    • Place of Presentation
      岡山 倉敷 倉敷市文芸館
    • Year and Date
      2014-06-08 – 2014-06-09
    • Related Report
      2014 Research-status Report
  • [Presentation] Analysis of HHV-6 TRS copy numbers in clinical isolates: Possibility of superinfection2013

    • Author(s)
      Yuri Kato, Masaru Ihira, Yuki Higashimoto, Yoshiki Kawamura, Masahiro Ohashi, Tetsushi Yoshikawa
    • Organizer
      8th International Conference on Human Herpesviruses 6 and 7
    • Place of Presentation
      Paris France.
    • Related Report
      2013 Research-status Report
  • [Presentation] Direct detection of HHV-6 DNA from serum by LAMP method using newly developed dry reagent2013

    • Author(s)
      Masaru Ihira, Yuki Higashimoto, Hiroko Sugiyama, Yoshihiko Enomoto,Yoshiki Kawamura, Masahiro Ohashi, Yoshizo Asano,Tetsushi Yoshikawa
    • Organizer
      8th International Conference on Human Herpesviruses 6 and 7
    • Place of Presentation
      Paris France.
    • Related Report
      2013 Research-status Report
  • [Presentation] Cytokine and chemokine response in the blood and cerebrospinal fluid of human herpesvirus 6B associated acute encephalopathy with biphasic seizures and late reduced diffusion2013

    • Author(s)
      Yoshiki Kawamura, Yuri Kato, Masahiro Ohashi, Masaru Ihira, Tetsushi Yoshikawa
    • Organizer
      8th International Conference on Human Herpesviruses 6 and 7
    • Place of Presentation
      Paris France.
    • Related Report
      2013 Research-status Report
  • [Presentation] Dual roles of telomeric repeats in chromosomally integrated human herpesvirus-62013

    • Author(s)
      Tamae Ohye, Hidehito Inagaki, Masaru Ihira, Yuki Higashimoto, Tetsushi Yoshikawa, Hiroki Kurahashi
    • Organizer
      8th International Conference on Human Herpesviruses 6 and 7
    • Place of Presentation
      Paris France.
    • Related Report
      2013 Research-status Report
  • [Presentation] Development of loop-mediated isothermal amplification (LAMP) method for detection of trans-renal Mycobacterium tuberculosis DNA in Urine2013

    • Author(s)
      M.Ihira, Chie Nakajima Yasuhiko Suzuki T. Yoshikawa and Y. Asano
    • Organizer
      US-Japan cooperative medical science program ;tuberculosis and leprosy panel Meeting
    • Place of Presentation
      Hokkaido Sapporo
    • Related Report
      2013 Research-status Report
  • [Presentation] LAMP 法によるMycobacterium tuberculosis特異的 rans-renal DNA検出

    • Author(s)
      井平 勝、中島千絵、浅野喜造、鈴木定彦、吉川哲史
    • Organizer
      第6回LAMP研究会
    • Place of Presentation
      東京
    • Related Report
      2013 Research-status Report

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Published: 2014-07-25   Modified: 2019-07-29  

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