Effect of substrate thermal resistance on space-domain microchannel fluorescent melting curve analysis.
In recent years, Fluorescent Melting Curve Analysis (FMCA) has become an almost ubiquitous feature of commercial quantitative PCR (qPCR) thermal cyclers. Here a micro-fluidic device is presented capable of performing FMCA within a microchannel. The device consists of modular thermally conductive blocks which can sandwich a microfluidic substrate. Opposing ends of the blocks are held at differing temperatures and a linear thermal gradient is generated along the microfluidic channel. Fluorescent measurements taken from a sample as it passes along the micro-fluidic channel permits fluorescent melting curves to be generated. In this study we measure DNA melting temperature from two plasmid fragments. The effects of flow velocity and ramp-rate are investigated, and measured melting curves are compared to those acquired from a commercially available PCR thermocycler.
Kinahan
David J
DJ
Stokes Bio Ltd., Henry St., Limerick, Ireland. david.kinahan@stokesbio.ie
Dalton
Tara M
TM
Davies
Mark R D
MR
D005453:Fluorescence; D006358:Hot Temperature; D046210:Microfluidic Analytical Techniques / Q000295:instrumentation* / Q000379:methods*; D010957:Plasmids / Q000737:chemistry*; D016133:Polymerase Chain Reaction / Q000295:instrumentation* / Q000379:methods*; D017422:Sequence Analysis, DNA / Q000295:instrumentation / Q000379:methods