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This thesis reports on the development of a fully integrated and automated microsystem consisting of low-cost, disposable plastic chips for DNA extraction and PCR amplification, combined with a reusable glass capillary array electrophoresis chip, which can be employed in a modular-based format for genetic analysis. In the thesis, DNA extraction is performed by adopting a filter paper-based method, followed by an "in-situ" PCR carried out directly in the same reaction chamber of the chip without elution. PCR products are then co-injected with sizing standards into separation channels for detection using a novel injection electrode. The entire process is automatically carried out by a custom-made compact control and detection instrument. The author thoroughly tests the system's performance and reliability by conducting rapid genetic screening of mutations on congenital hearing loss and pharmacogenetic typing of multiple warfarin-related single-nucleotide polymorphisms. The successfuldevelopment and operation of this microsystem establishes the feasibility of rapid "sample-in-answer-out" testing in routine clinical practice.
List of contents
Introduction.- The construction of a general platform for capillary electrophoresis.- Integrated module for automatic DNA extraction and amplification.- A fully-integrated genetic analysis system.- Conclusion and prospects for future work.
About the author
For internal review:
Dr. Bin Zhuang
Supervisor: Prof. Jing Cheng, School of Medicine, Tsinghua University
Current Affiliation: National Engineering Research Center for Beijing Biochip Technology, Capitalbio Corporation
Research Area: Biomedical engineering, microfluidic, nucleotide analysis, electrophoresis, biochemistry
Publications:
[1] Gan, W.P.,# Zhuang, B.,# Zhang, P.F., Han, J.P., Li, C.X., and Liu, P. (2014). A filter paper-based microdevice for low-cost, rapid, and automated DNA extraction and amplification from diverse sample types. Lab on a Chip 14, 3719-3728.
[2] Zhuang, B., # Gan, W.P., Wang, # S.Q., Han, J.P., Xiang, G.X., Li, C.X., Sun, J., Liu, P. (2015). A fully automated sample preparation microsystem for genetic testing of hereditary hearing loss using two-color multiplex allele-specific PCR. Analytical Chemistry, 87(2):1202-1209.
[3] Zhuang, B., # Han, J.P., # Xiang, G.X., Gan, W.P., Wang, S.Q., Wang, D., Wang, L., Sun, J., Li, C.X., Liu, P. (2016). A fully integrated and automated microsystem for rapid pharmacogenetic typing of multiple warfarin-related single-nucleotide polymorphisms. Lab on a Chip 16, 86-95.
Summary
This thesis reports on the development of a fully integrated and automated microsystem consisting of low-cost, disposable plastic chips for DNA extraction and PCR amplification, combined with a reusable glass capillary array electrophoresis chip, which can be employed in a modular-based format for genetic analysis. In the thesis, DNA extraction is performed by adopting a filter paper-based method, followed by an “in-situ” PCR carried out directly in the same reaction chamber of the chip without elution. PCR products are then co-injected with sizing standards into separation channels for detection using a novel injection electrode. The entire process is automatically carried out by a custom-made compact control and detection instrument. The author thoroughly tests the system’s performance and reliability by conducting rapid genetic screening of mutations on congenital hearing loss and pharmacogenetic typing of multiple warfarin-related single-nucleotide polymorphisms. The successfuldevelopment and operation of this microsystem establishes the feasibility of rapid “sample-in-answer-out” testing in routine clinical practice.
