Abstract
The polymerase chain reaction (PCR) for amplification of DNA has become a very useful tool in scientific research and analytical laboratories, yet conventional techniques are time-consuming, and the reagents are expensive. Miniaturization of this technique has the potential to drastically reduce amplification time and reagent consumption while simultaneously improving the efficiency of the reaction. Increasing the surface area-to-volume ratio using microfluidic reaction chambers allows homogeneous solution temperatures to be achieved much more rapidly than in conventional heating blocks. Employing infrared radiation to selectively heat the reaction solution can additionally reduce the time and energy needed for thermocycling; the reaction container is not heated and can even serve as a heat sink for enhancement of cooling. Microchip systems also provide the potential for fabrication of structures for additional processing steps directly in line with the PCR chamber. Not only can amplification be integrated with product separation and analysis, but sample preparation steps can also be incorporated prior to amplification. The ultimate goal is a miniature total-analysis-system with seamlessly coupled sample-in/answer-out capabilities that consumes very low volumes of reagents and drastically reduces the time for analysis. This chapter will focus on the materials and methods involved in simple straight-channel microchip PCR on glass substrates using non-contact thermocycling.
This is a preview of subscription content, log in via an institution.
Buying options
Tax calculation will be finalised at checkout
Purchases are for personal use only
Learn about institutional subscriptionsReferences
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.
Mai M., Grabs R., Barnes R. D., Vafiadis P., and Polychronakos C. (1998) Shortened PCR cycles in a conventional thermal cycler. Biotechniques 25, 208–210.
Wittwer C. T. and Garling D. J. (1991) Rapid cycle DNA amplification: time and temperature optimization. Biotechniques 10, 76–83.
Wilding P., Shoffner M. A., and Kricka6L. J. (1994) PCR in a silicon microstructure. Clin. Chem. 40, 1815–1818.
Wilding P., Shoffner M. A., Cheng J., Huichia G., and Kricka L. J. (1995) Thermal cycling and surface passivation of micromachined devices for PCR. Clin. Chem. 41, 1367, 1368.
Kopp M. U., Mello6A. J., and Manz A. (1998) Chemical amplification: continuousflow PCR on a chip. Science 280, 1046–1048.
Kricka L. J. and Wilding P. (2003) Microchip PCR. Anal. Bioanal. Chem. 377, 820–825.
Waters L. C., Jacobson S. C., Kroutchinina N., Khandurina J., Foote R. S., and Ramsey J. M. (1998) Microchip device for cell lysis, multiplex PCR amplification, and electrophoretic sizing. Anal. Chem. 70, 158–162.
Burns M. A., Mastrangelo C. H., Sammarco T. S., et al. (1996) Microfabricated structures for integrated DNA analysis. Proc. Natl. Acad. Sci. USA 93, 5556–5561.
Lagally, E T., Emrich C. A., and Mathies R. A. (2001) Fully integrated PCRcapillary electrophoresis microsystem for DNA analysis. Lab on a Chip 1, 102–107.
Koh C. G., Tan W., Zhao M. Q., Ricco A. J., and Fan Z. H. (2003) Integrating polymerase chain reaction, valving, and electrophoresis in a plastic device for bacterial detection. Anal. Chem. 75, 4591–4598.
Oda R. P., Strausbauch M. A., Huhmer A. F., et al. (1998) Infrared-mediated thermocycling for ultrafast polymerase chain reaction amplification of DNA. Anal. Chem. 70, 4361–4368.
Huhmer A. F. and Landers J. P. (2000) Noncontact infrared-mediated thermocycling for effective polymerase chain reaction amplification of DNA in nanoliter volumes. Anal. Chem. 72, 5507–5512.
Giordano B. C., Ferrance J., Swedberg S. Huhmer A. F., and Landers J. P. (2001) Polymerase chain reaction in polymeric microchips: DNA amplification in less than 240 seconds. Anal. Biochem. 291, 124–132.
Ferrance J. P., Wu Q., Giordano B., et al. (2003) Developments toward a complete micro-total analysis system for Duchenne muscular dystrophy diagnosis. Analytica Chimica Acta 500, 223–236.
Giordano B. C., Copeland E. R., and Landers J. P. (2001) Towards dynamic coating of glass microchip chambers for amplifying DNA via the polymerase chain reaction. Electrophoresis 22, 334–340.
Chiari, M., Cretich, M., Damin, F., Ceriotti, L., and Consonni, R. (2000) New adsorbed coatings for capillary electrophoresis. Electrophoresis 21, 909–916.
Author information
Authors and Affiliations
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2006 Humana Press Inc.
About this protocol
Cite this protocol
Easley, C.J., Legendre, L.A., Landers, J.P., Ferrance, J.P. (2006). Rapid DNA Amplification in Glass Microdevices. In: Henry, C.S. (eds) Microchip Capillary Electrophoresis. Methods in Molecular Biology, vol 339. Humana Press. https://doi.org/10.1385/1-59745-076-6:217
Download citation
DOI: https://doi.org/10.1385/1-59745-076-6:217
Publisher Name: Humana Press
Print ISBN: 978-1-58829-293-3
Online ISBN: 978-1-59745-076-8
eBook Packages: Springer Protocols