Development of a total integrated rotary genetic analysis microsystem for rapid and multiplex pathogen detection

Abstract

Pathogen detection to prevent the human epidemics or pandemics at early stages has emerged as a key issue in the fields of food industry, clinical diagnosis, and public health. Thus, the development of fast, sensitive, on-site pathogen detection methodologies has been intensively pursued. Among the methods for pathogen detection, genetic analysis has been strongly investigated due to high sensitivity, analytical robust-ness, low sample consumption, and high-throughput capability. Recently, microfluidic based genetic analysis garnered high attention due to the great advantages such as high detection sensitivity, rapid reaction speed, automation, and portability with a cost-effective manner. Thus, the development of total integrated microdevices for genetic analysis has been progressed for on-site pathogen detection with sample-in-answer-out capacity. However, the existing integrated microdevices require complicated chip fabrication, external pumps for fluid control, and well-trained experts for the operation of microdevice. In this thesis, the development of a novel genetic analysis system based on the centrifugal microfluidics, which have advantages including simple microchip design and fabrication, fully automated fluidic control without use of micropumps, microvalves, and high data reproducibility is presented. In Chapter 3, a novel of rotary reverse transcription-PCR (RT-PCR) system, called rotary PCR genetic analyzer was demonstrated for detecting influenza A virus subtypes (H1N1, H3N2, and H5N1). The rotary genetic analyzer consists of three parts including a disposable plastic PCR microchip, three thermal blocks equipped with resistive temperature detectors (RTD) for temperature control, and a stepper motor for precise rotation of the chip. The microchip is rotated on the thermal blocks from denature (95 °C) to annealing (58 °C) to extension (72 °C) for performance of the target gene amplification. The RT-PCR amplicons of influenza A viruses were produce...