Skip to main content

Polymerase Chain Reaction on Microchips

  • Protocol
Microfluidic Techniques

Part of the book series: Methods In Molecular Biology™ ((MIMB,volume 321))

Abstract

The polymerase chain reaction (PCR) provides an in vitro method for rapid enzymatic amplification of fragments of DNA. Microchip-based PCR devices (with reaction volumes from picoliters to microliters) have been realized using various combinations of silicon, glass, and/or plastic materials. Passivation of exposed surfaces in the microreactor is critical for successful PCR. Silicon and plastic surfaces can be passivated by silanization. With surface passivation, PCR can be performed efficiently and economically in chip-based microreactors. The reduced thermal mass of microchips allows for extremely fast temperature ramping. PCR protocols established for benchtop reactors may need to be adjusted accordingly when transferred to microchips. Here, we provide detailed protocols for microchip PCR including procedures for surface passivation and bonding of glass to silicon with ultraviolet curable glue, because both procedures have a major influence on the success or failure of the PCR.

This is a preview of subscription content, log in via an institution to check access.

Access this chapter

Protocol
USD 49.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD 84.99
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever
Softcover Book
USD 109.99
Price excludes VAT (USA)
  • Compact, lightweight edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info
Hardcover Book
USD 109.99
Price excludes VAT (USA)
  • Durable hardcover edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info

Tax calculation will be finalised at checkout

Purchases are for personal use only

Institutional subscriptions

References

  1. Gibbs, R. A. (1990) DNA amplification by the polymerase chain reaction. Anal. Chem. 62, 1202–1214.

    Article  PubMed  CAS  Google Scholar 

  2. Saiki, R. K., Scharf, S., Faloona, F., et al. (1985) Enzymatic amplification of betaglobin genomic sequences and restriction site analysis for diagnosis of sickle cell anemia. Science 230, 1350–1354.

    Article  PubMed  CAS  Google Scholar 

  3. Saiki, R. K., Chang, C. A., Levenson, C. H., et al. (1988) Diagnosis of sickle cell anemia and beta-thalassemia with enzymatically amplified DNA and nonradioactive allele-specific oligonucleotide probes. N. Engl. J. Med. 319, 537–541.

    Article  PubMed  CAS  Google Scholar 

  4. Bartlett, J. and Stirling, D. (2003) A short history of the polymerase chain reaction, in Methods in Molecular Biology: PCR Protocols, vol 226, 2nd ed. (Bartlett, J. and Stirling, D., eds.), Humana, Totowa, NJ, pp. 1–6.

    Chapter  Google Scholar 

  5. Kricka, L. J. and Wilding, P. (2003) Microchip PCR. Anal. Bioanal. Chem. 377, 820–825.

    Article  PubMed  CAS  Google Scholar 

  6. Reyes, D. R., Iossifidis, D., Auroux, P. A., and Manz, A. (2002) Micro total analysis systems. 1. Introduction, theory, and technology. Anal. Chem. 74, 2623–2636.

    Article  PubMed  CAS  Google Scholar 

  7. Liu, J., Enzelberger, M., and Quake, S. (2002) A nanoliter rotary device for polymerase chain reaction. Electrophoresis 23, 1531–1536.

    Article  PubMed  CAS  Google Scholar 

  8. Huber, M., Losert, D., Hiller, R., Harwanegg, C., Mueller, M. W., and Schmidt, W. M. (2001) Detection of single base alterations in genomic DNA by solid phase polymerase chain reaction on oligonucleotide microarrays. Anal. Biochem. 299, 24–30.

    Article  PubMed  CAS  Google Scholar 

  9. Auroux, P. A., Iossifidis, D., Reyes, D. R., and Manz, A. (2002) Micro total analysis systems. 2. Analytical standard operations and applications. Anal. Chem. 74, 2637–2652.

    Article  PubMed  CAS  Google Scholar 

  10. Vilkner, T., Janasek, D., and Manz, A. (2004) Micro total analysis systems: recent developments. Anal. Chem. 76, 3373–3385.

    Article  PubMed  CAS  Google Scholar 

  11. Carles, M., Lee, T., Moganti, S., et al. (2001) Chips and Qi: microcomponentbased analysis in traditional Chinese medicine. Fresenius J. Anal. Chem. 371, 190–194.

    Article  PubMed  CAS  Google Scholar 

  12. Dieffenbach, C. and Dveksler, G. (2003) PCR Primer: A Laboratory Manual, 2nd ed., Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY.

    Google Scholar 

  13. Innis, M., Gelfand, D., Sninsky, J., and White, T. (1990) PCR Protocols: A Guide to Methods and Applications, Academic, New York.

    Google Scholar 

  14. McPherson, M., Møller, S., Beynon, R., and Howe, C. (2000) PCR Basics: from Background to Bench, Springer Verlqag, New York, NY.

    Google Scholar 

  15. Shoffner, M. A., Cheng, J., Hvichia, G. E., Kricka, L. J., and Wilding, P. (1996) Chip PCR. I. Surface passivation of microfabricated silicon-glass chips for PCR. Nucleic Acids Res. 24, 375–379.

    Article  PubMed  CAS  Google Scholar 

  16. Lee, T. M., Hsing, I. M., Lao, A. I., and Carles, M. C. (2000) A miniaturized DNA amplifier: its application in traditional Chinese medicine. Anal. Chem. 72, 4242–4247.

    Article  PubMed  CAS  Google Scholar 

  17. Trau, D., Lee, T. M., Lao, A. I., Lenigk, R., et al. (2002) Genotyping on a complementary metal oxide semiconductor silicon polymerase chain reaction chip with integrated DNA microarray. Anal. Chem. 74, 3168–3173.

    Article  PubMed  CAS  Google Scholar 

  18. Woolley, A. T., Hadley, D., Landre, P., deMello, A. J., Mathies, R. A., and Northrup, M. A. (1996) Functional integration of PCR amplification and capillary electrophoresis in a microfabricated DNA analysis device. Anal. Chem. 68, 4081–4086.

    Article  PubMed  CAS  Google Scholar 

  19. Sucher, N. J., Deitcher, D. L., Baro, D. J., Warrick, R. M., and Guenther, E. (2000) Genes and channels: patch/voltage-clamp analysis and single-cell RT-PCR. Cell Tissue Res. 302, 295–307.

    Article  PubMed  CAS  Google Scholar 

  20. Sucher, N. J. and Deitcher, D. L. (1995) PCR and patch-clamp analysis of single neurons. Neuron 14, 1095–1100.

    Article  PubMed  CAS  Google Scholar 

  21. Rozen, S. and Skaletsky, H. (2000) Primer3 on the www for general users and for biologist programmers, in Bioinformatics Methods and Protocols: Methods in Molecular Biology (Krawetz, S. and Misener, S., eds.), Humana, Totowa, NJ, pp. 365–386.

    Google Scholar 

  22. Lagally, E. T., Medintz, I., and Mathies, R. A. (2001) Single-molecule DNA amplification and analysis in an integrated microfluidic device. Anal. Chem. 73, 565–570.

    Article  PubMed  CAS  Google Scholar 

Download references

Author information

Authors and Affiliations

Authors

Editor information

Editors and Affiliations

Rights and permissions

Reprints and permissions

Copyright information

© 2006 Humana Press Inc.

About this protocol

Cite this protocol

Carles, M.C., Sucher, N.J. (2006). Polymerase Chain Reaction on Microchips. In: Minteer, S.D. (eds) Microfluidic Techniques. Methods In Molecular Biology™, vol 321. Humana Press. https://doi.org/10.1385/1-59259-997-4:131

Download citation

  • DOI: https://doi.org/10.1385/1-59259-997-4:131

  • Publisher Name: Humana Press

  • Print ISBN: 978-1-58829-517-0

  • Online ISBN: 978-1-59259-997-4

  • eBook Packages: Springer Protocols

Publish with us

Policies and ethics