Written by: Karmella Haynes.
The course concluded with presentations from our 16 students about the work they did over the two weeks of lab training and research.
The TXTL projects (cell-free transcription-translation) were led by Julius Lucks and TA Melissa Takahashi. TXTL proved to be such a flexible system that almost every student had results to present from various in-course collaborations and explorations. Student presenters were Matt Munson, Ariel Hecht, Sarada Raghavan, Souvik Ghosh, Larry Kennedy, Clayton Coffman, Ileng Kumaran, Komal Singh, Maria Inda, Daniel Woodsworth, Katherine Brechun, and Taylor Canady. One sub-group used cell-free systems to switch on and off the expression of a gene using only RNA. Another group carried out validation assays to determine whether TXTL could be used as a controlled, reproducible, high-throughput platform to measure hundreds of genetic circuits. In order to speed up the Design, Build, Test cycle, one team combined Golden Gate DNA assembly for the building stage with TXTL for the testing stage. Another team tackled the challenge of reducing GFP reporter signal noise in TXTL during the inactivate state. One student integrated the work from her home lab to see if TXTL could support the engineering of a yellow fluorescent protein. Finally, the students presented the results of a mash-up project where they integrated CRISPR resources from the Mammalian Synthetic Biology and the Bacterial Engineering projects with TXTL, which we hope to continue after the course.
The Mammalian Synthetic Biology projects were headed by Karmella Haynes and Rene Davis. Student presenters were Larry Kennedy, Parijat Bhatnagar, Komal Singh, Ileng Kumaran, and Ashley Chessher. The group named their mCherry-expressing model tissue culture system “Emcherryitis” (a fictional disease) which was used to validate CRISPR as a tool for editing DNA in living cells. The goal was to use different combinations of the Cas9 protein and sequence-specific guide-RNAs to mutate and deactivate mCherry fluorescence. They identified two effective CRISPR plasmids and presented qPCR and SURVEYOR data to characterize the mutated DNA. They used an inducible ectopic chromatin system to determine whether chromatin interferes with CRISPR editing in human cells. They also designed and ran a trial of a CRISPR-editing scheme to improve a synthetic reporter by swapping luciferase for mCherry.
The Rapid Engineering of Bacteria projects were led by David Savage and Rayka Yokoo. Student presenters were Sarada Raghavan, Maria Inda, and Taylor Canady. The team used a violacein metabolism intermediate as a fluorescent signal to track synthesis in living bacteria. They used MAGE (multiplex automated genome engineering) to rapidly change genes in the bacteria and alter the fluorescence signal produced by the cells. The team also used a transposon knock-out library to hunt for genes that alter the behavior of the violacein-producing cells.
The Optogenetics projects were led by Jeff Tabor and Evan Olson. Student presenters were Katherine Brechun, Daniel Woodsworth, Ileng Kumaran, and Maria Inda. They used a system developed in Tabor’s lab to control the expression of a gene in bacteria using light instead of small molecules. Light has the advantages of acting independently of diffusion and being programmable (from a light-emitting device), and is not subject to dilution, degradation, or turnover. The light-driven gene expression system is developed so that anyone can use a computer algorithm for the specific light inputs that will generate a specific gene expression profile. The students reported their experiences, demonstrating the utility of the system.
The presentations concluded with a graduation ceremony for all sixteen of our fantastic students! Each received a signed certificate of completion from the instructors.
The instructors also gave special acknowledgement to the teaching assistants, Evan Olson, Rene Davis, Rayka Konoo, and Melissa Takahashi. Each received a beautiful 3-D printed, hand-painted, limited edition (4) unicorn trophy!