Biomedical Microdevices

, Volume 10, Issue 4, pp 539–546 | Cite as

Loop-mediated isothermal amplification of a single DNA molecule in polyacrylamide gel-based microchamber

  • Liza Lam
  • Shouichi Sakakihara
  • Koji Ishizuka
  • Shoji Takeuchi
  • Hideyuki F. Arata
  • Hiroyuki Fujita
  • Hiroyuki Noji
Article

Abstract

Loop-mediated isothermal amplification (LAMP) is an original nucleic acid amplification method established by Notomi et al. LAMP is performed under isothermal condition, employing only a basic reaction protocol and minimal supporting electronics. These requirements prove to be viable for exploring the avenues to down-scale this biological reaction for Lab-on-a-chip application. Hence here, we developed a novel technique for fluorescent imaging of LAMP at a single molecule level. The experiment was conducted in a polyacrylamide (PAA) gel-based microchamber where a single DNA template, freely suspended in a solution containing primers and polymerase was initially encapsulated. In order to activate the amplification reaction, a microheater regulated by an automatic computerized feedback system was used for localized heating. This microchamber-based approach for LAMP demonstrated the effective exploitation of minute amount of templates and primers, and the overall reduction in LAMP detection time. An average efficiency of 80% was evaluated for conducting DNA amplification after 50 min of incubation at 65°C. As the total time for reaction including detection can be completed in less than 1 h, this one-step, direct observation method displays the potential as a simple alternative to conventional techniques for genetic analysis and diagnosis in the clinical laboratory.

Keywords

DNA amplification Microchamber Polyacrylamide Single molecule level 

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Copyright information

© Springer Science+Business Media, LLC 2008

Authors and Affiliations

  • Liza Lam
    • 1
  • Shouichi Sakakihara
    • 1
  • Koji Ishizuka
    • 1
  • Shoji Takeuchi
    • 2
  • Hideyuki F. Arata
    • 2
  • Hiroyuki Fujita
    • 2
  • Hiroyuki Noji
    • 1
  1. 1.The Institute of Scientific and Industrial ResearchOsaka UniversityOsakaJapan
  2. 2.Center for International Research on MicroMechatronics, Institute of Industrial ScienceUniversity of TokyoTokyoJapan

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