Title page for ETD etd-10252011-122025

Type of Document Dissertation
Author Chen, Hui-Wen
URN etd-10252011-122025
Title Fully Integrated Microsystem for Bacterial Genotyping
Degree Doctor of Philosophy (Ph.D.)
Department Chemistry
Advisory Committee
Advisor Name Title
Soper, Steven A. Committee Chair
Crowe, William Committee Member
Hall, Randall Committee Member
Murray, Kermit K. Committee Member
Bollich, Patrick K. Dean's Representative
  • DNA analysis
  • microfluidics
Date of Defense 2011-08-16
Availability unrestricted
Methods for bacterial detection and identification has garnered renewed interest in

recent years due to the infections they may cause and the antimicrobial resistances

they can develop, the potential for bioterrorism threats and possible contamination of

food/water supplies. Therefore, the rapid, specific and accurate detection of pathogens

is crucial for the prevention of pathogen-related disease outbreaks and facilitating

disease management as well as the containment of suspected contaminated food

and/or water supplies. In this dissertation an integrated modular-based microfluidic

system composed of a fluidic cartridge and a control instrument has been developed for

bacterial pathogen detection. The integrated system can directly carry out the entire

molecular processing pipeline in a single disposable fluidic cartridge and can detect

sequence variations in selected genes to allow for the identification of the bacterial

species and even its strain. The unique aspect of this fluidic cartridge is its modular

format with a task-specific module interconnected to a fluidic motherboard to permit the

selection of a material appropriate for the given processing step(s). In addition, to

minimize the amount of finishing steps for assembling the fluidic cartridge, many of the

functional components were produced during the polymer molding step used to create

the fluidic network. The operation of the fluidic cartridge was provided by electronic,

mechanical, optical and hydraulic controls located off-chip and assembled into a small

footprint instrument. The fluidic cartridge was capable of performing cell lysis, solidphase

extraction of genomic DNA from the whole cell lysate, continuous flow PCR

amplification of specific gene fragments, continuous flow ligase detection reaction to

discriminate sequence variations and universal DNA array readout, which consisted of

DNA probes patterned onto a planar polymer waveguide for evanescent excitation. The

performance of the fluidic system was demonstrated through its successful application

to the genetic detection of bacterial pathogens, such as Escherichia coli O157:H7,

Salmonella, methicillin-resistant Staphylococcus aureus and multi-drug resistant

Mycobacterium tuberculosis, which are major threats for global heath. The modular

system, which could successfully identify several strains of bacteria in <40 min with

minimal human intervention and also perform strain identification, represents a

significant contribution to pathogen detection.

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