The polymerase chain reaction (PCR) is a manner of replicating specific sequences of deoxyribonucleic acid (DNA). The primary components involved in a typical reaction include free nucleotides, oligonucleotide primers, and a DNA polymerase. Primers define the region of the DNA strand to be copied, nucleotides serve as building blocks for the new strands of DNA and the polymerase acts as a cayalyst for DNA synthesis. The process is controlled by altering the temperature of the sample. By cycling through specified temperatures, it is possible to create millions of strands from just a single target strand of DNA. The simplicity of its technique has allowed for its integration within fields such as drug discovery, molecular archaeology, forensics, microbial detection, and most notably the diagnosis of both genetic and infectious diseases.
Following its introduction to the field of microfluidics, scientists and engineers have worked to exploit the advantages of this tandem including the ability to reduce reagent consumption, cost, and reaction time and in particular, improve portability and automation. With the integration of a miniature fluorescent detection module, these lab-on-a-chip (LOC) systems near the realization of a fully-integrated portable real-time PCR system useful for point-of-care testing of highly infectious agents.
My work in this area evaluates the use of inexpensive, disposable, microfluidic chips to detect infections agents. One of the projects involved the detection of methicillin-resistant Staphylococcus aureus (MRSA) that has undergone three forms of sample preparation: one containing purified genomic DNA, another containing the supernatant of a crude preparation using simple reagents, and a third through boiled culture preparation without any additional reagents. Polydimethylsiloxane (PDMS) microfluidic chips were fabricated by soft lithography and used with a miniature thermal cycler based on a thin resistive heater and a small fan to cycle through desired temperatures for the polymerase chain reaction (PCR). Fluorescent intensity measurements were taken at each cycle to monitor DNA replication and produce a standard curve. The details and results can be found in my thesis uploaded for the general public to Vanderbilt University's electronic thesis and dissertation database.
Following its introduction to the field of microfluidics, scientists and engineers have worked to exploit the advantages of this tandem including the ability to reduce reagent consumption, cost, and reaction time and in particular, improve portability and automation. With the integration of a miniature fluorescent detection module, these lab-on-a-chip (LOC) systems near the realization of a fully-integrated portable real-time PCR system useful for point-of-care testing of highly infectious agents.
My work in this area evaluates the use of inexpensive, disposable, microfluidic chips to detect infections agents. One of the projects involved the detection of methicillin-resistant Staphylococcus aureus (MRSA) that has undergone three forms of sample preparation: one containing purified genomic DNA, another containing the supernatant of a crude preparation using simple reagents, and a third through boiled culture preparation without any additional reagents. Polydimethylsiloxane (PDMS) microfluidic chips were fabricated by soft lithography and used with a miniature thermal cycler based on a thin resistive heater and a small fan to cycle through desired temperatures for the polymerase chain reaction (PCR). Fluorescent intensity measurements were taken at each cycle to monitor DNA replication and produce a standard curve. The details and results can be found in my thesis uploaded for the general public to Vanderbilt University's electronic thesis and dissertation database.
Design and Experimental Validation of a Miniature Real-time Polymerase
Chain Reaction Devices Using Disposable Microfluidic Chips
Master's Thesis written by Dustin House
Related journals and texts
 Encyclopedia of Microfluidics and NanofluidicsEditor-in-Chief: Dr. Dongqing Li This comprehensive encyclopedia provides easy access to information on all aspects of Microfluidics and Nanofluidics. With an A–Z format spanning over 500 entries, the encyclopedia is be a valuable resource for (analytical, organic, medicinal, physical, surface, and colloid) chemists and biochemists; molecular and cell biologists; geneticists; applied and industrial microbiologists; virologists; electronic, materials, chemical, mechanical, electrical engineers and bioengineers; pharmacists; bioprocess technologists; and upper-level undergraduate and graduate students in the field of microfluidics, nanofluidics and lab-on-a-chip devices. |
