Experimental Synthetic Biology, 2015

In this theme, students will learn the basics of the modern Synthetic Biology. In the morning sessions, they will attend basic synthetic biology concepts and methods lectures jointly with the Computational Synthetic Biology track, and in the afternoon sessions they will learn how to use experimental techniques common in synthetic and quantitative biology through demonstrations, hands-on tutorials, and group project wet lab work.

Lecturers

• Eva-Maria S. Collins, University of California, San Diego
• Suckjoon Jun, University of California, San Diego
• Nan Hao, University of California, San Diego
• Jeff M. Hasty, University of California, San Diego
• William Mather, Virginia Tech
• Stephen Mayfield, University of California, San Diego
• Justin Meyer, University of California, San Diego
• Brian Munsky, Colorado State University
• Gurol Suel, University of California, San Diego
• Lev S. Tsimring, University of California, San Diego
• Ruth Williams, University of California, San Diego
• Elizabeth Winzler, University of California, San Diego
• Roy Wollman, University of California, San Diego

Project Mentors

• Nan Hao
• Nick Csisery
• Meng Jin
• Yang Li
• Omar Din
• Ricky O'Laughlin
• Joydeb Sinha
• Yuan Zhao

Project Overview

Calorie restriction (CR) is the most robust environmental intervention that promotes longevity from yeast to mammals. However, the mechanism by which CR enhances longevity remains elusive. Using budding yeast S. cerevisiae as a model system, we will systematically characterize how CR affects the dynamics of abundance and subcellular localization of aging-related proteins in single cells over a long period of time. In particular, each student group will tag two selected aging-related genes with a genetically encoded fluorescent protein at their native loci, respectively. Microfluidics-incorporated time-lapse microscopy will be conducted to track the dynamic changes of these proteins in single cells in response to glucose limitation. Time-dependent fluorescence trajectories of individual cells will be further quantified and analyzed. Findings from this project will substantially advance our mechanistic understanding about the pro-longevity effects of CR.

Topics

• Basic bench techniques: Use of a pipettor, laboratory scale, incubator, gel station, waterbath. Instruction in media and reagent preparation, sterile technique, basic cell culture.
• Genetic circuit design: Instruction in the use of PCR primer design, PCR techniques, basic cloning techniques and troubleshooting in S. cerevisiae and E. coli. Quantitative analysis of genetic circuit behavior.
• Microfluidics: Instruction in device design using software tools, wafer fabrication using soft photolithography, device fabrication and QC, device testing.
• Microscopy: Instruction in Micromanager and Nikon Elements, setup of a microfluidic experiment with the dial-a-wave system, live cell imaging, proper setup and operation of a microfluidic experiment.
• Image analysis: Cell tracking and trajectory formation from microfluidic image data. Data analysis of cellular response to dynamic environments.

Schedule

• Experimental Synthetic Biology detailed program

Suggested Reading

[1] Gardner, T. S., Cantor, C. R. & Collins, J. J. Construction of a genetic toggle switch in Escherichia coli. Nature 403, 339–42 (2000).

[2] Elowitz, M. B. & Leibler, S. A synthetic oscillatory network of transcriptional regulators. Nature 403, 335–8 (2000).

[2] Hasty, J., McMillen, D. & Collins, J. J. Engineered gene circuits. Nature 420, 224–30 (2002).

[3] C. A. Voigt. Genetic parts to program bacteria. Curr. Opin. Biotech., 17(5) 548-557 (2006).

[4] P. E. M. Purnick & R. Weiss. The second wave of synthetic biology: from modules to systems. Nature Reviews Molecular Cell Biology 10, 410-422 (2009)

[5] J. Selimkhanov, J. Hasty, L. S. Tsimring. Recent advances in single-cell studies of gene regulation. Curr. Opin. Biotech., 23(1), 34-40 (2012).