SURF 2013: Synthetic biological circuit design implementing protein degradation in-vitro

2013 SURF project description

• Mentor: Richard Murray
• Co-mentor: Zachary Sun

Current methods of synthetic biological circuit design involve time-consuming cycles of plasmid construction, plasmid transformation, and circuit testing in-vivo. However, an E. coli crude extract developed by Shin and Noireaux (2010) shows properties emulating that of E. coli, enabling it to act as a model platform for prototyping circuits. Using this technology, we are currently developing a “biomolecular breadboard” for rapid prototyping of synthetic biological circuits entirely in-vitro.

Methods for protein degradation have been implemented in the biomolecular breadboard which seek to emulate protein concentration dilution by cell division. These methods allow for constructing circuits in-vitro previously impossible to probe biological phenomena. The summer project will focus on exploring circuits implementing protein degradation within the breadboard system, potentially drawing from in-vivo oscillators (see Elowitz and Libeler (2000) and Stricker et al. (2008)). Ultimately, the end goal is to answer an interesting biological question using a student-designed biological circuit. A significant portion of the work will be using molecular biological techniques such as cloning, gel electrophoresis, and cell culture. Synthetic biological circuit design will also be explored, as well as quantitative approaches.

Previous biological laboratory experience and proficiency with basic molecular biological techniques is a prerequisite. Experience using bioinformatics tools such as biopython and Geneious (or other sequence analysis software), and computational tools such as MATLAB is a plus but not necessary. Through this project, candidates can gain experience in the emerging field of synthetic biology, as well as in quantitative modeling and design, for later training in graduate school in the biological sciences and engineering.

References

Elowitz MB and Libeler S. A synthetic oscillatory network of transcriptional regulators. Nature 2008; 403: 335-338.

Stricker J, Cookson S, Bennett MR, Mather WH, Tsimring LS, Hasty J. A fast, robust, and tunable synthetic gene oscillator. Nature 2008; 456: 516-519.

Shin J and Noireaux V. Efficient cell-free expression with the endogenous E. Coli RNA polymerase and sigma factor 70. J. Biol. Eng. 2010; 4: 8.

Shin J and Noireaux V. Study of messenger RNA inactivation and protein degradation in an Escherichia coli cell-free expression system. J. Biol. Eng. 2010; 4: 9.

Shin J and Noireaux V. An E. coli cell-free expression toolbox: application to synthetic gene circuits and artificial cells. ACS Synth. Biol. 2012; 1: 29-41.

Siegal-Gaskins D, Noireaux V, Murray RM. Biomolecular response utilization in elementary cell-free circuits. Submitted to 2013 American Control Conference.