Date of Graduation

5-2015

Document Type

Thesis

Degree Name

Bachelor of Science in Biomedical Engineering

Degree Level

Undergraduate

Department

Biomedical Engineering

Advisor/Mentor

Barabote, Ravi

Abstract

Bacterial infection, or sepsis, places a disproportionately high burden on newborns in developing countries. This is due in part to a lack of diagnostic tools suitable for sustainable use in resource-limited nurseries. One potential vehicle for a new diagnostic assay is loop-mediated isothermal amplification (LAMP), a high-yield DNA amplification method. LAMP has been used to detect single genes from bacteria in blood serum samples to aid in sepsis diagnosis. While specific, this approach can only provide detection for one species at a time. LAMP could be adapted to detect a broader set of bacteria, while retaining a degree of specificity that allows clinicians to begin directed antimicrobial therapy. Described herein is the successful design of a novel group of oligonucleotide LAMP primer sets that specifically bind to regions of the 16s rRNA gene of four bacterial Orders. These regions lie on the transition area between sequences that are highly conserved and those that are hypervariable. This allows each primer set to be specific for one Order of bacteria. When primers bind, amplification occurs, and the large quantities of DNA produced could be detected using a fluorescent indicator. The four separate LAMP reactions could be multiplexed on a microfluidic chip to provide clinicians with a two-step sepsis-diagnosis technique that could give a result in only one hour. Future studies should examine fluorescent indicators, reaction multiplexing, and specificity of primers to recognize more species.

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