PhD Research

Genetic Engineering Yeast.

Since the summer of 2020, I have been working in the Ian Wheeldon lab at UC Riverside. From producing cannabinoids in yeast to designing “smart yeast” with molecular sensors, the general theme of my projects seek to genetically engineer yeast to produce novel chemicals and develop new genetic engineering tools.

Molecular Sensors

Sensors are a critical component in many circuits. The same holds true for biological circuits. Originally from the plant kingdom, the PYR1-HAB1 system provides a selective and sensitive molecular sensor circuit when tied to a DNA activation and binding domain. The PYR1 component can easily be mutated to accept new ligands. My work has shown that this sensor can be engineered to be selective between chemical categories like natural and synthetic cannabinoids, PFAS, organophosphates, and substances of abuse with nanomolar range sensitivity.

Genome-Wide CRISPR Screens

CRISPR Cas9 genome-wide screens enable rapid identification of gene targets for metabolic engineering. Our lab have developed these tools in non-conventional yeast to, in a single flask, obtain every possible single-gene knockout in Y. lipolytica, K. marxianus, or K. phafii. In Y. lipolytica, we developed an optimized, compact guide RNA library that targets 99% of genes. With this library, I was able to screen the yeast for gene knockouts that would improve tolerance/growth on alternative carbon sources like acetate, fatty acids, and long-chain hydrocarbons. These gene discoveries allow for more efficient utilization of low-cost/waste substrates for industrial production of valuable biochemicals from yeast.

Smart Yeast

By combining genome-wide CRISPR screening with PYR1 biosensor technology, we will be able to create mutant “smart yeast” that “know” how much chemical they are producing. I have built a strain of K. marxianus that produces a class of flavor and fragrance molecules called terpenes. By integrating a PYR1 biosensor that senses terpenes, my yeast “know” how much product they are making, thus allowing me to select for mutations that improve terpene production. By applying a previously developed genome-wide CRISPR screen for K. marxianus, I am able to rapidly identify non-obvious genes that improve terpene production.