BME Graduate Student Seminar (ZOOM): Hamsa Gowda and Ziwei Zhong

Seminar via Zoom link: https://uci.zoom.us/j/99821164165

Hamsa Gowda: Development of a Portable Pathogen Analysis System for Water Monitoring using a Centrifugal Microfluidics Platform

Abstract: Waterborne diseases cause millions of deaths worldwide, especially in developing communities. The detection of waterborne pathogens is a critical step for the selection of water treatment processes that will lead to the prevention of disease transmission. Unfortunately, analyzing water samples is a lengthy and laborious process that requires 1) collecting and transporting large volumes of samples (>1 L) to a centralized lab, 2) removing extraneous contaminants through filtration and concentration methods and 3) using cultivation or molecular techniques to detect the pathogen of interest. This lengthy “sample-to-answer” process significantly delays risk mitigation actions, and subsequently exposes those using the water for daily necessities to pathogen infection. In this study, we have developed a microfluidic-based portable pathogen analysis system (PPAS) that can detect bacterial contaminations in a point-of-sample setting. Specifically, a centrifugal microfluidic platform is adopted and designed to seamlessly integrate sample loading, genomic DNA extraction, fluid metering, reagent mixing, digital droplet isothermal amplification and image processing into one system. The centrifugal disc functions as a single-step bacterial detection “lab” without the need for intermediate vial-to-vial sample transfer, minimizing hands-on time, sample loss and potential contamination. The disc is loaded in a portable box-like enclosure (called PPAS cube) that includes an electric motor to drive the rotation of the disc, infrared lamps to heat the amplification reaction chamber, and a camera to capture the digital images. The cube is driven by a Raspberry Pi 3, and a touchscreen graphical user interface enables the user to start, stop and monitor the steps of the assay.  To demonstrate the capabilities of the system, we have shown the fluidic integration and detection of E. faecalis, a common waterborne pathogen, in culture samples within 45 minutes.

Ziwei Zhong: Engineering an Orthogonal Replication System to Study Protein Evolution

Abstract: Directed evolution is a powerful tool that has enabled the development of novel biological drugs and catalysts, as well as elucidated mechanisms of protein evolution. In a process analogous to natural evolution, genes of interest are subjected to multiple rounds of mutagenesis and selection to improve a desired trait, such as stability, activity or binding affinity. Traditionally, directed evolution has relied on multiple rounds of mutation generation with error-prone PCR or DNA shuffling, subsequent transformation into bacteria or yeast for protein production and selection, and finally isolation of the best performers to repeat the process again. However, this process is highly labor-intensive and time-consuming, thus limiting the scope of possible experiments and targets. Our lab has recently developed OrthoRep, an orthogonal replication system in yeast capable of mutating genes of interest at rates 100,000 greater than genomic mutation rates. As a result, genes of interest can be continuously and rapidly diversified in vivo, enabling high-throughput directed evolution of proteins that are capable of traversing long mutational pathways. I will present some of the technical improvements that I have developed for OrthoRep, including completely automating the evolution process, as well as using OrthoRep to explore the fitness landscape of proteins and the driving forces of protein evolution.