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DNA Purification on Microfluidic Devices With a Focus on Large Volume, Forensic Biological Samples

Reedy, Carmen Reneé
Thesis/Dissertation; Online
Reedy, Carmen Reneé
Landers, James
Harrison, Ian
Allen, Ralph
Venton, Jill
Demas, James
The development and application of microfluidic sample preparation methods to a wide range of sample types, including large-volume forensic samples undergoing genetic analysis, has the potential to greatly benefit the forensic and clinical communities. The design, development and optimization of a microfluidic solid phase extraction method applicable to the purification of DNA from forensic biological samples (vrSPE) obtained in large volumes is demonstrated. Illustration of the range of samples this method can handle, the successful application to environmentally-degraded DNA samples, mitochondrial DNA from blood and blood stains, and genomic DNA from bone are also shown. Integration of vrSPE with a secondary, orthogonal, purification method is demonstrated to be advantageous for the removal of the PCR inhibitors (e.g., indigo dye), and to outperform low volume microfluidic SPE (SPE) and vrSPE in the removal of that inhibitor. The development of a microfluidic, forensic genetic analysis system, both in a modular and integrated form, is detailed. The modular system described, used three microfluidic devices for SPE, polymerase chain reaction (PCR), and microchip electrophoresis (ME), while the integrated system used a single device for both SPE and PCR. Each method was used for the successful, forensic STR analysis of buccal swab lysate. Finally, an exploratory excursion into the development of a PMMA (Plexiglas) microfluidic device for DNA purification is shown, with a focus on device bonding and surface modification. This work provides the next step towards developing a single-use, genetic analysis microdevice. Note: Abstract extracted from PDF text
University of Virginia, Department of Chemistry, PHD, 2011
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