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Fluorescent Melting Curve Analysis Compatible With a Flowing Polymerase Chain Reactor

[+] Author Affiliations
Tara M. Dalton, David J. Kinahan, Mark R. Davies

University of Limerick

Paper No. IMECE2005-80954, pp. 31-36; 6 pages
doi:10.1115/IMECE2005-80954
From:
  • ASME 2005 International Mechanical Engineering Congress and Exposition
  • Advances in Bioengineering
  • Orlando, Florida, USA, November 5 – 11, 2005
  • Conference Sponsors: Bioengineering Division
  • ISBN: 0-7918-4213-4 | eISBN: 0-7918-3769-6
  • Copyright © 2005 by ASME

abstract

A primary tool for analysing PCR product is the Fluorescent Melting Curve Analysis (FMCA). The temperature at which a double helix DNA strand denatures depends both on its length and base pair composition. Accurate measurement of this melting temperature using fluorescence allows estimations be made regarding DNA product length and composition. Current progress in development of PCR thermal cyclers has been primarily aimed at micro-channel based flowing devices. This paper addresses the challenges associated with performing FMCA analysis which is compatible with the output from a flowing PCR thermocycler. Two PCR products of significantly different lengths and base pair composition are compared using space domain FMCA. Results allow for differentiation of the PCR product, and compare favourably with results from a commercial thermal cycler. The successful application of FMCA within a channel shows its potential for use in high throughput flow based total analysis systems (μTAS).

Copyright © 2005 by ASME
Topics: Melting , Chain

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