MEC Seminar Proteome Research Lab 2003. 5. 20 Summarized by Park, Ji-Yoon

Integrated Microsystem of Isothermal Amplification Integrated Microsystem of Isothermal Amplification of DNA and Electrophoresis on a Microfabricated Plof DNA and Electrophoresis on a Microfabricated Plastic Chip for Detection of Specific Gene and Analysastic Chip for Detection of Specific Gene and Analys

is of Genetic Materialsis of Genetic Materials

MEC Seminar MEC Seminar

Proteome Research LabProteome Research Lab

2003. 5. 20 2003. 5. 20

Summarized by Park, Ji-Yoon Summarized by Park, Ji-Yoon

Proceedings of the uTAS 2002 SymposiumProceedings of the uTAS 2002 Symposium, Vol. 1, pp 215-217, Vol. 1, pp 215-217

2002. 11. 27 2002. 11. 27 Proteome Research Lab, http://gene.hanyang.ac.kr/ Proteome Research Lab, http://gene.hanyang.ac.kr/

Continuous-Flow PCR on ChipContinuous-Flow PCR on Chip

ScienceScience, 280, 1046-1048(1998), 280, 1046-1048(1998)

Fig 1. Schematic of a chip for flow through PCRFig 1. Schematic of a chip for flow through PCR

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A Miniature Integrated Device for Automated Multiple A Miniature Integrated Device for Automated Multiple Genetic AssaysGenetic Assays

Nucleic Acids Research, Nucleic Acids Research, 28, e60 (2000) 28, e60 (2000)

Fig 4. An integrated device that automatically performs a multistep HIV genotyping assayFig 4. An integrated device that automatically performs a multistep HIV genotyping assay

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High Sensitivity PCR Assay in Plastic Micro Reactors High Sensitivity PCR Assay in Plastic Micro Reactors

Lab on a Chip, Lab on a Chip, 2(4), 179-187(2002)2(4), 179-187(2002)

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Infrared-mediated Thermocycling for DNA AmplificatiInfrared-mediated Thermocycling for DNA Amplification and Electrophoretic Separation on an Integrated Micron and Electrophoretic Separation on an Integrated Micr

orchip Deviceorchip Device


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Fig 2. Microchip and thermocycling profileFig 2. Microchip and thermocycling profile


DNA Amplification and Hybridization Assays in IntegrDNA Amplification and Hybridization Assays in Integrated Plastic Monolithic Devicesated Plastic Monolithic Devices


Fig 3. Monolithic integrated polycarbonatedFig 3. Monolithic integrated polycarbonated DNA assay device DNA assay device

Anal. Chem

Anal. Chem. 74, 3063-3070 (2002)

. 74, 3063-3070 (2002)

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On-chip integration of isothermal PCR and electrophoresis On-chip integration of isothermal PCR and electrophoresis Loop-mediated isothermal amplification(LAMP) Loop-mediated isothermal amplification(LAMP)

Novel DNA amplification method developed by Eiken Chemical Co., Ltd.Novel DNA amplification method developed by Eiken Chemical Co., Ltd. By using 4 different primers specifically designed to recognize six distinct regBy using 4 different primers specifically designed to recognize six distinct reg

ions on the target gene ions on the target gene Performed at a constant temperature using a strand displacement reaction. Performed at a constant temperature using a strand displacement reaction.

Advantages Advantages High amplification efficiency( 10High amplification efficiency( 1099-10-101010 times in 15-60 minutes) times in 15-60 minutes) High specificity High specificity The potential to replace PCRThe potential to replace PCR

» simplicity, rapidity, specificity and cost-effectiveness » simplicity, rapidity, specificity and cost-effectiveness Avoid the thermal cycling Avoid the thermal cycling

Analysis Analysis by the integrated microchip electrophoresis by the integrated microchip electrophoresis by the naked eyes by adding fluorescent dye like SYBR Green I by the naked eyes by adding fluorescent dye like SYBR Green I

AbstractAbstract

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Loop-mediated Isothermal Amplification Loop-mediated Isothermal Amplification (LAMP)(LAMP)

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RAMRAM(Ramplification Amplification Method)(Ramplification Amplification Method)

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Chip Device and ElectropherogramChip Device and Electropherogram

Integrated microchip for DNA amplificIntegrated microchip for DNA amplification and CE ation and CE

Reaction conditionReaction condition RAMP; 5- 10 ul volume, RAMP; 5- 10 ul volume,

50-65°C for 10-30min 50-65°C for 10-30min Micro-CEMicro-CE

- Reproducibility(Fig. 4) - Reproducibility(Fig. 4)

- Detection time; 10min (Fig 5)- Detection time; 10min (Fig 5)

- Visualization(Fig 6) - Visualization(Fig 6)

Fig 1. Structure of a PMMA chip andFig 1. Structure of a PMMA chip and device and electropherogram device and electropherogram

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SYBR Green™ ISYBR Green™ I

The dye (ex; EtBr)The dye (ex; EtBr)

Binds preferentially to ds DNA & quantitate the amount of double-stranBinds preferentially to ds DNA & quantitate the amount of double-stranded product ded product

Stable to the extremes of temperature required for PCR reactionStable to the extremes of temperature required for PCR reaction

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The Reproducibility and DetectionThe Reproducibility and Detection

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ConclusionsConclusions

Integrated microsystem Integrated microsystem Useful for fast, sensitive, and simple detection of specific sequence of Useful for fast, sensitive, and simple detection of specific sequence of

DNA DNA ApplicationApplication

- O-157 - O-157 E.coliE.coli, , B. anthracisB. anthracis, , SalmonellaSalmonella and other bacteria in the and other bacteria in the environment, food, animal, human blood, etc. environment, food, animal, human blood, etc.

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Further StudyFurther Study

Development of DNA Computing Chip Development of DNA Computing Chip DNA hybridization, ligation, PCR, CE DNA hybridization, ligation, PCR, CE the uTAS 2003 Symposiumthe uTAS 2003 Symposium

Application of DNA Computing Application of DNA Computing Diagnosis, Gene expression, Environmental Checker etc… Diagnosis, Gene expression, Environmental Checker etc…

uTAS 2002 Paper uTAS 2002 Paper To access the latest papers published in LOC please seeTo access the latest papers published in LOC please see http:// http://www.rsc.org/is/journals/current/loc/lcadvarts.htmwww.rsc.org/is/journals/current/loc/lcadvarts.htm For Advance articles For Advance articles

http://www.rsc.org/is/journals/current/loc/lc002003.htmhttp://www.rsc.org/is/journals/current/loc/lc002003.htm The general website is The general website is www.rsc.orgwww.rsc.org/loc/loc

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MEC Seminar Proteome Research Lab 2003. 5. 20 Summarized by Park, Ji-Yoon

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