Ribocomputing devices for sophisticated in vivo logic computation

ABSTRACT

Synthetic biology aims to create functional devices, systems, and organisms with novel and useful functions taking advantage of engineering principles applied to biology. Despite great progress over the last decade, an underlying problem in synthetic biology remains the limited number of high-performance, modular, composable parts. A potential route to solve parts bottleneck problem in synthetic biology utilizes the programmability of nucleic acids inspired by molecular programming approaches that have demonstrated complex biomolecular circuits evaluating logic expressions in test tubes. Using a library of de-novo-designed toehold switches with orthogonality and modular composability, we demonstrate how toehold switches can be incorporated into decision-making RNA networks termed ribocomputing devices to rapidly evaluate complex logic in living cells. We have successfully demonstrated a 4-input AND gate, a 6-input OR gate, and a 12-input expression in disjunctive normal form in E. coli. The compact encoding of ribocomputing system using a library of modular parts is amenable to aggressive scale-up towards complex control of in vivo circuitry towards autonomous behaviors and biomedical applications.

References

1. Chappell, J., Takahashi, M.K., and Lucks, J.B. (2015). Creating small transcription activating RNAs. Nature Chemical Biology 11, 214--220.
2. Daniel, R., Rubens, J.R., Sarpeshkar, R., and Lu, T.K. (2013). Synthetic analog computation in living cells. Nature 497, 619--623.
3. Elowitz, M.B., and Leibler, S. (2000). A synthetic oscillatory network of transcriptional regulators. Nature 403, 335--338.
4. Gardner, T.S., Cantor, C.R., and Collins, J.J. (2000). Construction of a genetic toggle switch in Escherichia coli. Nature 403, 339--342.
5. Green, A.A., Silver, P.A., Collins, J.J., and Yin, P. (2014). Toehold Switches: De-Novo-Designed Regulators of Gene Expression. Cell 159, 925--939.
6. Isaacs, F.J., Dwyer, D.J., Ding, C.M., Pervouchine, D.D., Cantor, C.R., and Collins, J.J. (2004). Engineered riboregulators enable post-transcriptional control of gene expression. Nature Biotechnology 22, 841--847.
7. Moon, T.S., Lou, C., Tamsir, A., Stanton, B.C., and Voigt, C.A. (2012). Genetic programs constructed from layered logic gates in single cells. Nature 491, 249--253.
8. Nielsen, A.A.K., Der, B.S., Shin, J., Vaidyanathan, P., Paralanov, V., Strychalski, E.A., Ross, D., Densmore, D., and Voigt, C.A. (2016). Genetic circuit design automation. Science 352.
9. Pardee, K., Green, A.A., Ferrante, T., Cameron, D.E., DaleyKeyser, A., Yin, P., and Collins, J.J. (2014). Paper-Based Synthetic Gene Networks. Cell 159, 940--954.

Index Terms

• Applied computing

  • Life and medical sciences

    • Computational biology

    • Genetics

    • Systems biology