Three-dimensional on-chip continuous-flow polymerase chain reaction employing a single heater |
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Authors: | Wenming?Wu Email author" target="_blank">Nae?Yoon?LeeEmail author |
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Institution: | (1) Division of BioNano Technology and College of BioNano Technology, Kyungwon University, San 65, Bokjeong-dong, Sujeong-gu, Seongnam, Gyeonggi-do, 461–701, South Korea;(2) Present address: BioNanotechnology Research Center, Korea Research Institute of Bioscience and Biotechnology (KRIBB), Daejeon, 305–600, South Korea; |
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Abstract: | Multi-step temperature control in a polymerase chain reaction (PCR) is a limiting factor in device miniaturization and portability.
In this study, we propose the fabrication of a three-dimensional (3D) microdevice employing a single heater to minimize temperature
control required for an on-chip continuous-flow PCR as well as the overall footprint by stacking the device in multi-layers.
Two poly(dimethylsiloxane) (PDMS) layers with differing thicknesses are vertically stacked with their microchannel-engraved
sides facing down. Through-holes are made in the thicker PDMS layer, which is sandwiched between a glass substrate at the
bottom and the thinner PDMS layer at the top. In this way, a fluidic conduit is realized in a 3D configuration. The assembled
3D microdevice is then placed onto a heater glass-side down. The interface of the two PDMS layers displays a relatively lower
temperature than that of the PDMS and glass layers due to the low thermal conductivity of the PDMS and its physical distance
from the heater. The denaturation temperature can be controlled by adjusting the temperature of the heater, while the annealing/extension
temperature can be controlled automatically by molding the thicker bottom PDMS layer into the appropriate thickness calculated
using a numerical derivation proposed in this study. In this way, a cumbersome temperature measurement step is eliminated.
DNA amplification was successfully carried out using the proposed 3D fluidic microdevice, and the intensity of the resulting
amplicon was comparable to that obtained using a thermal cycler. This novel concept of adopting a single heating source greatly
simplifies the temperature control issue present in an on-chip continuous-flow PCR. It also allows the use of a commercialized
hot plate as a potential heat source, paving the way for device miniaturization and portability in a highly cost-effective
manner. In this study, a simple and facile technique to make arrays of through-holes for the fluidic interconnection inside
a 3D channel configuration is also addressed. |
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