Nanodroplet real-time PCR system with laser assisted heating.
We report the successful application of low-power (approximately 30 mW) laser radiation as an optical heating source for high-speed real-time polymerase chain reaction (PCR) amplification of DNA in nanoliter droplets dispersed in an oil phase. Light provides the heating, temperature measurement, and Taqman real-time readout in nanoliter droplets on a disposable plastic substrate. A selective heating scheme using an infrared laser appears ideal for driving PCR because it heats only the droplet, not the oil or plastic substrate, providing fast heating and completing the 40 cycles of PCR in 370 seconds. No microheaters or microfluidic circuitry were deposited on the substrate, and PCR was performed in one droplet without affecting neighboring droplets. The assay performance was quantitative and its amplification efficiency was comparable to that of a commercial instrument.
Kim
Hanyoup
H
Molecular Physics Laboratory, SRI International, Menlo Park, California 94025, USA.
Dixit
Sanhita
S
Green
Christopher J
CJ
Faris
Gregory W
GW
D004247:DNA / Q000032:analysis / Q000235:genetics* / Q000528:radiation effects; D004867:Equipment Design; D005784:Gene Amplification; D006361:Heating / Q000295:instrumentation; D007259:Infrared Rays; D007834:Lasers; D008027:Light; D046210:Microfluidic Analytical Techniques / Q000295:instrumentation; D008904:Miniaturization; D016133:Polymerase Chain Reaction / Q000295:instrumentation / Q000379:methods*; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000295:instrumentation; D013696:Temperature; D013816:Thermodynamics
D004247:DNA