Download The 3rd Research on Theorem Proving

Integrated Microsystem of Isothermal Amplification
of DNA and Electrophoresis on a Microfabricated
Plastic Chip for Detection of Specific Gene and
Analysis of Genetic Materials
Proceedings of the uTAS 2002 Symposium, Vol. 1, pp 215-217
MEC Seminar
Proteome Research Lab
2003. 5. 20
Summarized by Park, Ji-Yoon
Continuous-Flow PCR on Chip
Science, 280, 1046-1048(1998)
Fig 1. Schematic of a chip for flow through PCR
2002. 11. 27
Proteome Research Lab, http://gene.hanyang.ac.kr/
7
A Miniature Integrated Device for Automated Multiple
Genetic Assays
Nucleic Acids Research, 28, e60 (2000)
Fig 4. An integrated device that automatically performs a multistep HIV genotyping assay
2002. 11. 27
Proteome Research Lab, http://gene.hanyang.ac.kr/
8
High Sensitivity PCR Assay in Plastic Micro Reactors
Lab on a Chip, 2(4), 179-187(2002)
2002. 11. 27
Proteome Research Lab, http://gene.hanyang.ac.kr/
9
Infrared-mediated Thermocycling for DNA
Amplification and Electrophoretic Separation on an
Integrated Microrchip Device
Proceedings of the uTAS 2002 Symposium, Vol. 1, pp 193-194
Fig 2. Microchip and thermocycling profile
2002. 11. 27
Proteome Research Lab, http://gene.hanyang.ac.kr/
10
DNA Amplification and Hybridization Assays in
Integrated Plastic Monolithic Devices
Proceedings of the uTAS 2002 Symposium, Vol. 1, pp 169-171
Fig 3. Monolithic integrated polycarbonated
DNA assay device
2002. 11. 27
Proteome Research Lab, http://gene.hanyang.ac.kr/
11
Abstract




On-chip integration of isothermal PCR and electrophoresis
Loop-mediated isothermal amplification(LAMP)
 Novel DNA amplification method developed by Eiken Chemical Co., Ltd.
 By using 4 different primers specifically designed to recognize six distinct
regions on the target gene
 Performed at a constant temperature using a strand displacement reaction.
Advantages
 High amplification efficiency( 109-1010 times in 15-60 minutes)
 High specificity
 The potential to replace PCR
» simplicity, rapidity, specificity and cost-effectiveness
 Avoid the thermal cycling
Analysis
 by the integrated microchip electrophoresis
 by the naked eyes by adding fluorescent dye like SYBR Green I
2002. 11. 27
Proteome Research Lab, http://gene.hanyang.ac.kr/
12
Loop-mediated Isothermal Amplification
(LAMP)
2002. 11. 27
Proteome Research Lab, http://gene.hanyang.ac.kr/
13
RAM(Ramplification Amplification Method)
2002. 11. 27
Proteome Research Lab, http://gene.hanyang.ac.kr/
14
Chip Device and Electropherogram
Fig 1. Structure of a PMMA chip and
device and electropherogram
2002. 11. 27

Integrated microchip for DNA
amplification and CE

Reaction condition
 RAMP; 5- 10 ul volume,
50-65°C for 10-30min
 Micro-CE
- Reproducibility(Fig. 4)
- Detection time; 10min (Fig 5)
- Visualization(Fig 6)
Proteome Research Lab, http://gene.hanyang.ac.kr/
15
SYBR Green™ I

The dye (ex; EtBr)

Binds preferentially to ds DNA & quantitate the amount of
double-stranded product

Stable to the extremes of temperature required for PCR reaction
2002. 11. 27
Proteome Research Lab, http://gene.hanyang.ac.kr/
6
The Reproducibility and Detection
2002. 11. 27
Proteome Research Lab, http://gene.hanyang.ac.kr/
16
Conclusions

Integrated microsystem


Useful for fast, sensitive, and simple detection of specific sequence of
DNA
Application
- O-157 E.coli, B. anthracis, Salmonella and other bacteria in the
environment, food, animal, human blood, etc.
2002. 11. 27
Proteome Research Lab, http://gene.hanyang.ac.kr/
17
Further Study

Development of DNA Computing Chip
 DNA hybridization, ligation, PCR, CE
 the uTAS 2003 Symposium

Application of DNA Computing
 Diagnosis, Gene expression, Environmental Checker etc…

uTAS 2002 Paper
 To access the latest papers published in LOC please see
http://www.rsc.org/is/journals/current/loc/lcadvarts.htm
 For Advance articles
http://www.rsc.org/is/journals/current/loc/lc002003.htm
 The general website is www.rsc.org/loc
2002. 11. 27
Proteome Research Lab, http://gene.hanyang.ac.kr/
18