Thermal Design, Analysis And Optimization Of Silicon-Based Micro Polymerase Chain Reaction Chip

Polymerase Chain Reaction (PCR) is a very important molecular biological method. A wide variety of DNA/RNA molecules can be amplified by this enzyme catalysis reaction and thereby enriched for the further analysis. In the past decades, micro PCR chip is one of the hotspots in the development of micro systems for the biological and clinical applications. In the present dissertation, thermal analysis, design and optimization of silicon-based micro PCR chips were given, and the silicon inhibition effects on nucleic acids amplification was studied experimentally. Numerical simulation and lumped heat capacity analysis were carried out to study the thermal performances of the micro chamber PCR chip and micro continuous-flow PCR chip. Effects of the chip geometries, materials and boundary conditions on the thermal cyclings of the PCR chips were investigated. The characteristics of the constant-temperature control in the chips were also analyzed. Optimizations were suggested based on the numerical and analytical results. Based on the studies of thermal performances of the micro chamber PCR chip and micro continuous-flow PCR chip, a novel thermal cycling model, named "heating/cooling –constant temperature"hybrid model was put forwards. And an original micro PCR chip, droplet-based micro oscillating-flow PCR chip, was fabricated by silicon microfabrication technique to realize the hybrid thermal cycling model. In the novel PCR chip, a droplet of the PCR mixture flew through three temperature zones in an oscillating model to realize the denaturation, annealing and extension processes. HPV-DNA was amplified by the present chip system, and the results demonstrated that the droplet-based micro oscillating-flow PCR chip can achieve fast and correct nucleic acids amplification, and the time needed for the chip PCR to finish the amplification is about 1/9 of that required by the conventional instrument. A lumped heat capacity model of the droplet-based micro oscillating flow PCR chip has been built to optimize the chip geometries and the heat sink temperature in the operation. Thermal performance of the optimized chip was numerically simulated, and the results indicated that an ultra-fast heating and cooling rates were achieved with very small temperature non-uniformities. Parameters used in the constant-temperature control were also optimized by a coupled simulation of the chip thermal cycling and the control system. In order to quantificationally analyze the inhibition effects of the native silicon oxide and the untreated silicon on the PCR, silicon nano-particles with different oxidized degrees were added into the PCR mixtures. A real-time PCR was carried out to study the amplification performance of these PCR mixtures. Mechanisms of the inhibition phenomena were also analyzed based on the experimental results.