Source: https://www.nature.com/articles/nmeth.2926?error=cookies_not_supported&code=26ed09d3-5ec2-4ca7-a8e3-4a7e9f84aa27
Timestamp: 2019-04-25 14:23:13+00:00

Document:
Cells navigate environments, communicate and build complex patterns by initiating gene expression in response to specific signals. Engineers seek to harness this capability to program cells to perform tasks or create chemicals and materials that match the complexity seen in nature. This Review describes new tools that aid the construction of genetic circuits. Circuit dynamics can be influenced by the choice of regulators and changed with expression 'tuning knobs'. We collate the failure modes encountered when assembling circuits, quantify their impact on performance and review mitigation efforts. Finally, we discuss the constraints that arise from circuits having to operate within a living cell. Collectively, better tools, well-characterized parts and a comprehensive understanding of how to compose circuits are leading to a breakthrough in the ability to program living cells for advanced applications, from living therapeutics to the atomic manufacturing of functional materials.
Dahl, R.H. et al. Engineering dynamic pathway regulation using stress-response promoters . Nat. Biotechnol. 31, 1039–1046 (2013).
Moser, F. et al. Genetic circuit performance under conditions relevant for industrial bioreactors . ACS Synth. Biol. 1, 555–564 (2012).
Holtz, W.J. & Keasling, J.D. Engineering static and dynamic control of synthetic pathways . Cell 140, 19–23 (2010).
Anesiadis, N., Kobayashi, H., Cluett, W.R. & Mahadevan, R. Analysis and design of a genetic circuit for dynamic metabolic engineering . ACS Synth. Biol. 2, 442–452 (2013).
Zhang, F. & Keasling, J. Biosensors and their applications in microbial metabolic engineering . Trends Microbiol. 19, 323–329 (2011).
Dietrich, J.A., Shis, D.L., Alikhani, A. & Keasling, J.D. Transcription factor-based screens and synthetic selections for microbial small-molecule biosynthesis . ACS Synth. Biol. 2, 47–58 (2013).
Schendzielorz, G. et al. Taking control over control: use of product sensing in single cells to remove flux control at key enzymes in biosynthesis pathways . ACS Synth. Biol. 3, 21–29 (2014).
Zhang, F., Carothers, J.M. & Keasling, J.D. Design of a dynamic sensor-regulator system for production of chemicals and fuels derived from fatty acids . Nat. Biotechnol. 30, 354–359 (2012).
Yi, T.-M., Huang, Y., Simon, M.I. & Doyle, J. Robust perfect adaptation in bacterial chemotaxis through integral feedback control . Proc. Natl. Acad. Sci. USA 97, 4649–4653 (2000).
Krishnanathan, K., Anderson, S.R., Billings, S.A. & Kadirkamanathan, V. A data-driven framework for identifying nonlinear dynamic models of genetic parts . ACS Synth. Biol. 1, 375–384 (2012).
Carbonell, P., Parutto, P., Baudier, C., Junot, C. & Faulon, J.-L. Retropath: automated pipeline for embedded metabolic circuits . ACS Synth. Biol. 10.1021/sb4001273 (4 October 2013).
Adams, B.L. et al. Evolved quorum sensing regulator, LsrR, for altered switching functions . ACS Synth. Biol. 10.1021/sb400068z (10 October 2013).
Umeyama, T., Okada, S. & Ito, T. Synthetic gene circuit-mediated monitoring of endogenous metabolites: identification of GAL11 as a novel multicopy enhancer of S-adenosylmethionine level in yeast . ACS Synth. Biol. 2, 425–430 (2013).
Stapleton, J.A. et al. Feedback control of protein expression in mammalian cells by tunable synthetic translational inhibition . ACS Synth. Biol. 1, 83–88 (2012).
Liu, D., Xiao, Y., Evans, B.S. & Zhang, F. Negative feedback regulation of fatty acid production based on a malonyl-CoA sensor-actuator . ACS Synth. Biol. 10.1021/sb400158w (30 December 2013).
Siedler, S. et al. SoxR as a single-cell biosensor for NADPH-consuming enzymes in Escherichia coli . ACS Synth. Biol. 3, 41–47 (2014).
Medema, M.H., Breitling, R., Bovenberg, R. & Takano, E. Exploiting plug-and-play synthetic biology for drug discovery and production in microorganisms . Nat. Rev. Microbiol. 9, 131–137 (2011).
Fischbach, M. & Voigt, C.A. Prokaryotic gene clusters: a rich toolbox for synthetic biology . Biotechnol. J. 5, 1277–1296 (2010).
Frasch, H.-J., Medema, M.H., Takano, E. & Breitling, R. Design-based re-engineering of biosynthetic gene clusters: plug-and-play in practice . Curr. Opin. Biotechnol. 24, 1144–1150 (2013).
Shao, Z. et al. Refactoring the silent spectinabilin gene cluster using a plug-and-play scaffold . ACS Synth. Biol. 2, 662–669 (2013).
Oßwald, C. et al. Modular construction of a functional artificial epothilone polyketide pathway . ACS Synth. Biol. 10.1021/sb300080t (25 October 2012).
Steidler, L. et al. Treatment of murine colitis by Lactococcus lactis secreting interleukin-10 . Science 289, 1352–1355 (2000).
Anderson, J.C., Clarke, E.J., Arkin, A.P. & Voigt, C.A. Environmentally controlled invasion of cancer cells by engineered bacteria . J. Mol. Biol. 355, 619–627 (2006).
Ruder, W.C., Lu, T. & Collins, J.J. Synthetic biology moving into the clinic . Science 333, 1248–1252 (2011).
Motta, J.-P. et al. Food-grade bacteria expressing elafin protect against inflammation and restore colon homeostasis . Sci. Transl. Med. 4, 158ra144 (2012).
Wang, S., Kong, Q. & Curtiss, R. III. New technologies in developing recombinant attenuated Salmonella vaccine vectors . Microb. Pathog. 58, 17–28 (2013).
Huh, J.H., Kittleson, J.T., Arkin, A.P. & Anderson, J.C. Modular design of a synthetic payload delivery device . ACS Synth. Biol. 2, 418–424 (2013).
Gupta, S., Bram, E.E. & Weiss, R. Genetically programmable pathogen sense and destroy . ACS Synth. Biol. 2, 715–723 (2013).
Hwang, I.Y. et al. Reprogramming microbes to be pathogen-seeking killers . ACS Synth. Biol. 10.1021/sb400077j (10 September 2013).
Prindle, A. et al. Genetic circuits in Salmonella typhimurium . ACS Synth. Biol. 1, 458–464 (2012).
Volzing, K., Borrero, J., Sadowsky, M.J. & Kaznessis, Y.N. Antimicrobial peptides targeting gram-negative pathogens, produced and delivered by lactic acid bacteria . ACS Synth. Biol. 2, 643–650 (2013).
Hasty, J. Engineered microbes for therapeutic applications . ACS Synth. Biol. 1, 438–439 (2012).
Danino, T., Lo, J., Prindle, A., Hasty, J. & Bhatia, S.N. In vivo gene expression dynamics of tumor-targeted bacteria . ACS Synth. Biol. 1, 465–470 (2012).
Archer, E.J., Robinson, A.B. & Süel, G.M. Engineered E. coli that detect and respond to gut inflammation through nitric oxide sensing . ACS Synth. Biol. 1, 451–457 (2012).
Antunes, M.S. et al. Programmable ligand detection system in plants through a synthetic signal transduction pathway . PLoS ONE 6, e16292 (2011).
Widmaier, D.M. et al. Engineering the Salmonella type III secretion system to export spider silk monomers . Mol. Syst. Biol. 5, 309 (2009).
Bernhardt, K. et al. New tools for self-organized pattern formation . BMC Syst. Biol. 1 (suppl. 1), S10 (2007).
Xia, X.-X. et al. Native-sized recombinant spider silk protein produced in metabolically engineered Escherichia coli results in a strong fiber . Proc. Natl. Acad. Sci. USA 107, 14059–14063 (2010).
Widmaier, D.M. & Voigt, C.A. Quantification of the physiochemical constraints on the export of spider silk proteins by Salmonella type III secretion . Microb. Cell Fact. 9, 78 (2010).
Aquea, F. et al. A molecular framework for the inhibition of Arabidopsis root growth in response to boron toxicity . Plant Cell Environ. 35, 719–734 (2012).
Antunes, M.S. et al. A synthetic de-greening gene circuit provides a reporting system that is remotely detectable and has a re-set capacity . Plant Biotechnol. J. 4, 605–622 (2006).
Purnick, P.E.M. & Weiss, R. The second wave of synthetic biology: from modules to systems . Nat. Rev. Mol. Cell Biol. 10, 410–422 (2009).
Lewis, N.E., Nagarajan, H. & Palsson, B.O. Constraining the metabolic genotype-phenotype relationship using a phylogeny of in silico methods . Nat. Rev. Microbiol. 10, 291–305 (2012).
Weiss, R. Cellular Computation and Communications Using Engineered Genetic Regulatory Networks. PhD thesis, MIT (2001).
Salis, H.M., Mirsky, E.A. & Voigt, C.A. Automated design of synthetic ribosome binding sites to control protein expression . Nat. Biotechnol. 27, 946–950 (2009).
Cambray, G. et al. Measurement and modeling of intrinsic transcription terminators . Nucleic Acids Res. 41, 5139–5148 (2013).
Rodrigo, G. & Jaramillo, A. AutoBioCAD: full biodesign automation of genetic circuits . ACS Synth. Biol. 2, 230–236 (2013).
Voigt, C.A. Genetic parts to program bacteria . Curr. Opin. Biotechnol. 17, 548–557 (2006).
Yokobayashi, Y., Weiss, R. & Arnold, F.H. Directed evolution of a genetic circuit . Proc. Natl. Acad. Sci. USA 99, 16587–16591 (2002).
Ellefson, J.W. et al. Directed evolution of genetic parts and circuits by compartmentalized partnered replication . Nat. Biotechnol. 32, 97–101 (2014).
Moon, T.S., Lou, C., Tamsir, A., Stanton, B.C. & Voigt, C.A. Genetic programs constructed from layered logic gates in single cells . Nature 491, 249–253 (2012).
Haseltine, E.L. & Arnold, F.H. Synthetic gene circuits: design with directed evolution . Annu. Rev. Biophys. Biomol. Struct. 36, 1–19 (2007).
Collins, C.H., Arnold, F.H. & Leadbetter, J.R. Directed evolution of Vibrio fischeri LuxR for increased sensitivity to a broad spectrum of acyl-homoserine lactones . Mol. Microbiol. 55, 712–723 (2005).
Sleight, S.C. & Sauro, H.M. Randomized BioBrick assembly: a novel DNA assembly method for randomizing and optimizing genetic circuits and metabolic pathways . ACS Synth. Biol. 2, 506–518 (2013).
Shong, J. & Collins, C.H. Engineering the esaR promoter for tunable quorum sensing-dependent gene expression . ACS Synth. Biol. 2, 568–575 (2013).
Balagaddé, F.K., You, L., Hansen, C.L., Arnold, F.H. & Quake, S.R. Long-term monitoring of bacteria undergoing programmed population control in a microchemostat . Science 309, 137–140 (2005).
Cardinale, S. & Arkin, A.P. Contextualizing context for synthetic biology – identifying causes of failure of synthetic biological systems . Biotechnol. J. 7, 856–866 (2012).
Engler, C., Gruetzner, R., Kandzia, R. & Marillonnet, S. Golden Gate shuffling: a one-pot DNA shuffling method based on type IIs restriction enzymes . PLoS ONE 4, e5553 (2009).
Hillson, N.J., Rosengarten, R.D. & Keasling, J.D. j5 DNA assembly design automation software . ACS Synth. Biol. 1, 14–21 (2012).
Leguia, M., Brophy, J.A., Densmore, D., Asante, A. & Anderson, J.C. 2ab assembly: a methodology for automatable, high-throughput assembly of standard biological parts . J. Biol. Eng. 7, 2 (2013).
de Kok, S. et al. Rapid and reliable DNA assembly via ligase cycling reaction . ACS Synth. Biol. 3, 97–106 (2014).
Paetzold, B., Carolis, C., Ferrar, T., Serrano, L. & Lluch-Senar, M. In situ overlap and sequence synthesis during DNA assembly . ACS Synth. Biol. 2, 750–755 (2013).
Clancy, K. & Voigt, C.A. Programming cells: towards an automated 'genetic compiler' . Curr. Opin. Biotechnol. 21, 572–581 (2010).
Stanton, B.C. et al. Genomic mining of prokaryotic repressors for orthogonal logic gates . Nat. Chem. Biol. 10, 99–105 (2014).
Endy, D. Foundations for engineering biology . Nature 438, 449–453 (2005).
Khalil, A.S. & Collins, J.J. Synthetic biology: applications come of age . Nat. Rev. Genet. 11, 367–379 (2010).
Liang, J.C., Bloom, R.J. & Smolke, C.D. Engineering biological systems with synthetic RNA molecules . Mol. Cell 43, 915–926 (2011).
Lim, W.A. Designing customized cell signalling circuits . Nat. Rev. Mol. Cell Biol. 11, 393–403 (2010).
Weber, W. & Fussenegger, M. Synthetic gene networks in mammalian cells . Curr. Opin. Biotechnol. 21, 690–696 (2010).
Liu, W., Yuan, J.S. & Stewart, C.N. Jr. Advanced genetic tools for plant biotechnology . Nat. Rev. Genet. 14, 781–793 (2013).
Bikard, D. et al. Programmable repression and activation of bacterial gene expression using an engineered CRISPR-Cas system . Nucleic Acids Res. 41, 7429–7437 (2013).
Liu, C.C. et al. An adaptor from translational to transcriptional control enables predictable assembly of complex regulation . Nat. Methods 9, 1088–1094 (2012).
Mutalik, V.K., Qi, L., Guimaraes, J.C., Lucks, J.B. & Arkin, A.P. Rationally designed families of orthogonal RNA regulators of translation . Nat. Chem. Biol. 8, 447–454 (2012).
Beerli, R.R. & Barbas, C.F. III. Engineering polydactyl zinc-finger transcription factors . Nat. Biotechnol. 20, 135–141 (2002).
Garg, A., Lohmueller, J.J., Silver, P.A. & Armel, T.Z. Engineering synthetic TAL effectors with orthogonal target sites . Nucleic Acids Res. 40, 7584–7595 (2012).
Moscou, M.J. & Bogdanove, A.J. A simple cipher governs DNA recognition by TAL effectors . Science 326, 1501 (2009).
Takeda, Y., Folkmanis, A. & Echols, H. Cro regulatory protein specified by bacteriophage λ . J. Biol. Chem. 252, 6177–6183 (1977).
Ptashne, M. & Hopkins, N. The operators controlled by the lambda phage repressor . Proc. Natl. Acad. Sci. USA 60, 1282–1287 (1968).
Zhan, J. et al. Develop reusable and combinable designs for transcriptional logic gates . Mol. Syst. Biol. 6, 388 (2010).
Elowitz, M.B. & Leibler, S. A synthetic oscillatory network of transcriptional regulators . Nature 403, 335–338 (2000).
Guet, C.C., Elowitz, M.B., Hsing, W. & Leibler, S. Combinatorial synthesis of genetic networks . Science 296, 1466–1470 (2002).
Hasty, J., Dolnik, M., Rottschäfer, V. & Collins, J.J. Synthetic gene network for entraining and amplifying cellular oscillations . Phys. Rev. Lett. 88, 148101 (2002).
Hooshangi, S., Thiberge, S. & Weiss, R. Ultrasensitivity and noise propagation in a synthetic transcriptional cascade . Proc. Natl. Acad. Sci. USA 102, 3581–3586 (2005).
Gaber, R. et al. Designable DNA-binding domains enable construction of logic circuits in mammalian cells . Nat. Chem. Biol. 10, 203–208 (2014).
Lohmueller, J.J., Armel, T.Z. & Silver, P.A. A tunable zinc finger-based framework for Boolean logic computation in mammalian cells . Nucleic Acids Res. 40, 5180–5187 (2012).
Peacock, R.W.S., Sullivan, K.A. & Wang, C.L. Tetracycline-regulated expression implemented through transcriptional activation combined with proximal and distal repression . ACS Synth. Biol. 1, 156–162 (2012).
Mercer, A.C., Gaj, T., Sirk, S.J., Lamb, B.M. & Barbas, C.F. III. Regulation of endogenous human gene expression by ligand-inducible TALE transcription factors . ACS Synth. Biol. 10.1021/sb400114p (19 November 2013).
Purcell, O., Peccoud, J. & Lu, T.K. Rule-based design of synthetic transcription factors in eukaryotes . ACS Synth. Biol. 10.1021/sb400134k (12 December 2013).
Lienert, F. et al. Two- and three-input TALE-based AND logic computation in embryonic stem cells . Nucleic Acids Res. 41, 9967–9975 (2013).
Rhodius, V.A. et al. Design of orthogonal genetic switches based on a crosstalk map of σs, anti-σs, and promoters . Mol. Syst. Biol. 9, 702 (2013).
Wang, B., Kitney, R.I., Joly, N. & Buck, M. Engineering modular and orthogonal genetic logic gates for robust digital-like synthetic biology . Nat. Commun. 2, 508 (2011).
Anderson, J.C., Voigt, C.A. & Arkin, A.P. Environmental signal integration by a modular AND gate . Mol. Syst. Biol. 3, 133 (2007).
Daniel, R., Rubens, J.R., Sarpeshkar, R. & Lu, T.K. Synthetic analog computation in living cells . Nature 497, 619–623 (2013).
Buchler, N.E., Gerland, U. & Hwa, T. On schemes of combinatorial transcription logic . Proc. Natl. Acad. Sci. USA 100, 5136–5141 (2003).
Calles, B. & de Lorenzo, V. Expanding the boolean logic of the prokaryotic transcription factor XylR by functionalization of permissive sites with a protease-target sequence . ACS Synth. Biol. 2, 594–603 (2013).
Ramalingam, K.I., Tomshine, J.R., Maynard, J.A. & Kaznessis, Y.N. Forward engineering of synthetic bio-logical AND gates . Biochem. Eng. J. 47, 38–47 (2009).
Lou, C. et al. Synthesizing a novel genetic sequential logic circuit: a push-on push-off switch . Mol. Syst. Biol. 6, 350 (2010).
Ausländer, S., Ausländer, D., Müller, M., Wieland, M. & Fussenegger, M. Programmable single-cell mammalian biocomputers . Nature 487, 123–127 (2012).
Shis, D.L. & Bennett, M.R. Library of synthetic transcriptional AND gates built with split T7 RNA polymerase mutants . Proc. Natl. Acad. Sci. USA 110, 5028–5033 (2013).
Basu, S., Mehreja, R., Thiberge, S., Chen, M.-T. & Weiss, R. Spatiotemporal control of gene expression with pulse-generating networks . Proc. Natl. Acad. Sci. USA 101, 6355–6360 (2004).
Chen, D. & Arkin, A.P. Sequestration-based bistability enables tuning of the switching boundaries and design of a latch . Mol. Syst. Biol. 8, 620 (2012).
Atkinson, M.R., Savageau, M.A., Myers, J.T. & Ninfa, A.J. Development of genetic circuitry exhibiting toggle switch or oscillatory behavior in Escherichia coli . Cell 113, 597–607 (2003).
Fung, E. et al. A synthetic gene–metabolic oscillator . Nature 435, 118–122 (2005).
Tigges, M., Dénervaud, N., Greber, D., Stelling, J. & Fussenegger, M. A synthetic low-frequency mammalian oscillator . Nucleic Acids Res. 38, 2702–2711 (2010).
Argos, P. et al. The integrase family of site-specific recombinases: regional similarities and global diversity . EMBO J. 5, 433–440 (1986).
Gopaul, D.N. & Van Duyne, G.D. Structure and mechanism in site-specific recombination . Curr. Opin. Struct. Biol. 9, 14–20 (1999).
Ham, T.S., Lee, S.K., Keasling, J.D. & Arkin, A.P. A tightly regulated inducible expression system utilizing the fim inversion recombination switch . Biotechnol. Bioeng. 94, 1–4 (2006).
Moon, T.S. et al. Construction of a genetic multiplexer to toggle between chemosensory pathways in Escherichia coli . J. Mol. Biol. 406, 215–227 (2011).
Bonnet, J., Yin, P., Ortiz, M.E., Subsoontorn, P. & Endy, D. Amplifying genetic logic gates . Science 340, 599–603 (2013).
Siuti, P., Yazbek, J. & Lu, T.K. Synthetic circuits integrating logic and memory in living cells . Nat. Biotechnol. 31, 448–452 (2013).
Sorek, R., Lawrence, C.M. & Wiedenheft, B. CRISPR-mediated adaptive immune systems in bacteria and archaea . Annu. Rev. Biochem. 82, 237–266 (2013).
Sashital, D.G., Wiedenheft, B. & Doudna, J.A. Mechanism of foreign DNA selection in a bacterial adaptive immune system . Mol. Cell 46, 606–615 (2012).
Farzadfard, F., Perli, S.D. & Lu, T.K. Tunable and multifunctional eukaryotic transcription factors based on CRISPR/Cas . ACS Synth. Biol. 2, 604–613 (2013).
Gilbert, L.A. et al. CRISPR-mediated modular RNA-guided regulation of transcription in eukaryotes . Cell 154, 442–451 (2013).
Maeder, M.L. et al. CRISPR RNA-guided activation of endogenous human genes . Nat. Methods 10, 977–979 (2013).
Perez-Pinera, P. et al. RNA-guided gene activation by CRISPR-Cas9–based transcription factors . Nat. Methods 10, 973–976 (2013).
Esvelt, K.M. et al. Orthogonal Cas9 proteins for RNA-guided gene regulation and editing . Nat. Methods 10, 1116–1121 (2013).
Larson, M.H. et al. CRISPR interference (CRISPRi) for sequence-specific control of gene expression . Nat. Protoc. 8, 2180–2196 (2013).
Gossen, M., Bonin, A.L. & Bujard, H. Control of gene activity in higher eukaryotic cells by prokaryotic regulatory elements . Trends Biochem. Sci. 18, 471–475 (1993).
Sternberg, S.H., Redding, S., Jinek, M., Greene, E.C. & Doudna, J.A. DNA interrogation by the CRISPR RNA-guided endonuclease Cas9 . Nature 507, 62–67 (2014).
Del Vecchio, D., Ninfa, A.J. & Sontag, E.D. Modular cell biology: retroactivity and insulation . Mol. Syst. Biol. 4, 161 (2008).
Deltcheva, E. et al. CRISPR RNA maturation by trans-encoded small RNA and host factor RNase III . Nature 471, 602–607 (2011).
Simons, R.W. & Kleckner, N. Translational control of IS10 transposition . Cell 34, 683–691 (1983).
Kittle, J.D., Simons, R.W., Lee, J. & Kleckner, N. Insertion sequence IS10 anti-sense pairing initiates by an interaction between the 5′ end of the target RNA and a loop in the anti-sense RNA . J. Mol. Biol. 210, 561–572 (1989).
Ma, C. & Simons, R.W. The IS10 antisense RNA blocks ribosome binding at the transposase translation initiation site . EMBO J. 9, 1267–1274 (1990).
Qi, L., Lucks, J.B., Liu, C.C., Mutalik, V.K. & Arkin, A.P. Engineering naturally occurring trans-acting non-coding RNAs to sense molecular signals . Nucleic Acids Res. 40, 5775–5786 (2012).
Liu, C.C., Qi, L., Yanofsky, C. & Arkin, A.P. Regulation of transcription by unnatural amino acids . Nat. Biotechnol. 29, 164–168 (2011).
Callura, J.M., Dwyer, D.J., Isaacs, F.J., Cantor, C.R. & Collins, J.J. Tracking, tuning, and terminating microbial physiology using synthetic riboregulators . Proc. Natl. Acad. Sci. USA 107, 15898–15903 (2010).
Nielsen, A.A., Segall-Shapiro, T.H. & Voigt, C.A. Advances in genetic circuit design: novel biochemistries, deep part mining, and precision gene expression . Curr. Opin. Chem. Biol. 17, 878–892 (2013).
Bintu, L. et al. Transcriptional regulation by the numbers: models . Curr. Opin. Genet. Dev. 15, 116–124 (2005).
Bintu, L. et al. Transcriptional regulation by the numbers: applications . Curr. Opin. Genet. Dev. 15, 125–135 (2005).
Voigt, C.A., Wolf, D.M. & Arkin, A.P. The Bacillus subtilis sin operon: an evolvable network motif . Genetics 169, 1187–1202 (2005).
Strogatz, S.H. Nonlinear Dynamics and Chaos: With Application to Physics, Biology, Chemistry, and Engineering (Westview, 2000).
Ang, J., Harris, E., Hussey, B.J., Kil, R. & McMillen, D.R. Tuning response curves for synthetic biology . ACS Synth. Biol. 2, 547–567 (2013).
Gottesman, S. Proteases and their targets in Escherichia coli . Annu. Rev. Genet. 30, 465–506 (1996).
Naryshkin, N., Revyakin, A., Kim, Y., Mekler, V. & Ebright, R.H. Structural organization of the RNA polymerase-promoter open complex . Cell 101, 601–611 (2000).
Kittleson, J.T., Cheung, S. & Anderson, J.C. Rapid optimization of gene dosage in E. coli using DIAL strains . J. Biol. Eng. 5, 10 (2011).
Cox, R.S. III., Surette, M.G. & Elowitz, M.B. Programming gene expression with combinatorial promoters . Mol. Syst. Biol. 3, 145 (2007).
Chen, S. et al. Automated design of genetic toggle switches with predetermined bistability . ACS Synth. Biol. 1, 284–290 (2012).
Koshland, D.E., Goldbeter, A. & Stock, J.B. Amplification and adaptation in regulatory and sensory systems . Science 217, 220–225 (1982).
Vilar, J.M. & Saiz, L. DNA looping in gene regulation: from the assembly of macromolecular complexes to the control of transcriptional noise . Curr. Opin. Genet. Dev. 15, 136–144 (2005).
Johnson, S., Lindén, M. & Phillips, R. Sequence dependence of transcription factor-mediated DNA looping . Nucleic Acids Res. 40, 7728–7738 (2012).
Legewie, S., Dienst, D., Wilde, A., Herzel, H. & Axmann, I.M. Small RNAs establish delays and temporal thresholds in gene expression . Biophys. J. 95, 3232–3238 (2008).
Buchler, N.E. & Cross, F.R. Protein sequestration generates a flexible ultrasensitive response in a genetic network . Mol. Syst. Biol. 5, 272 (2009).
Lu, M.S., Mauser, J.F. & Prehoda, K.E. Ultrasensitive synthetic protein regulatory networks using mixed decoys . ACS Synth. Biol. 1, 65–72 (2012).
Lee, T.-H. & Maheshri, N. A regulatory role for repeated decoy transcription factor binding sites in target gene expression . Mol. Syst. Biol. 8, 576 (2012).
Cookson, N.A. et al. Queueing up for enzymatic processing: correlated signaling through coupled degradation . Mol. Syst. Biol. 7, 561 (2011).
Shen, J., Liu, Z., Zheng, W., Xu, F. & Chen, L. Oscillatory dynamics in a simple gene regulatory network mediated by small RNAs . Physica A 388, 2995–3000 (2009).
Geyer, P.K. The role of insulator elements in defining domains of gene expression . Curr. Opin. Genet. Dev. 7, 242–248 (1997).
Lou, C., Stanton, B., Chen, Y.-J., Munsky, B. & Voigt, C.A. Ribozyme-based insulator parts buffer synthetic circuits from genetic context . Nat. Biotechnol. 30, 1137–1142 (2012).
Jeong, W. & Kang, C. Start site selection at lacUV5 promoter affected by the sequence context around the initiation sites . Nucleic Acids Res. 22, 4667–4672 (1994).
Walker, K.A. & Osuna, R. Factors affecting start site selection at the Escherichia coli fis promoter . J. Bacteriol. 184, 4783–4791 (2002).
Kudla, G., Murray, A.W., Tollervey, D. & Plotkin, J.B. Coding-sequence determinants of gene expression in Escherichia coli . Science 324, 255–258 (2009).
Goodman, D.B., Church, G.M. & Kosuri, S. Causes and effects of N-terminal codon bias in bacterial genes . Science 342, 475–479 (2013).
Kosuri, S. et al. Composability of regulatory sequences controlling transcription and translation in Escherichia coli . Proc. Natl. Acad. Sci. USA 110, 14024–14029 (2013).
Qi, L., Haurwitz, R.E., Shao, W., Doudna, J.A. & Arkin, A.P. RNA processing enables predictable programming of gene expression . Nat. Biotechnol. 30, 1002–1006 (2012).
Chen, Y.-J. et al. Characterization of 582 natural and synthetic terminators and quantification of their design constraints . Nat. Methods 10, 659–664 (2013).
Yao, A.I. et al. Promoter element arising from the fusion of standard BioBrick parts . ACS Synth. Biol. 2, 111–120 (2013).
Villalobos, A., Ness, J.E., Gustafsson, C., Minshull, J. & Govindarajan, S. Gene Designer: a synthetic biology tool for constructing artificial DNA segments . BMC Bioinformatics 7, 285 (2006).
Rhodius, V.A., Mutalik, V.K. & Gross, C.A. Predicting the strength of UP-elements and full-length E. coli σE promoters . Nucleic Acids Res. 40, 2907–2924 (2012).
Brewster, R.C., Jones, D.L. & Phillips, R. Tuning promoter strength through RNA polymerase binding site design in Escherichia coli . PLoS Comput. Biol. 8, e1002811 (2012).
Weller, K. & Recknagel, R.-D. Promoter strength prediction based on occurrence frequencies of consensus patterns . J. Theor. Biol. 171, 355–359 (1994).
Seo, S.W. et al. Predictive design of mRNA translation initiation region to control prokaryotic translation efficiency . Metab. Eng. 15, 67–74 (2013).
de Hoon, M.J.L., Makita, Y., Nakai, K. & Miyano, S. Prediction of transcriptional terminators in Bacillus subtilis and related species . PLoS Comput. Biol. 1, e25 (2005).
Lesnik, E.A. et al. Prediction of rho-independent transcriptional terminators in Escherichia coli . Nucleic Acids Res. 29, 3583–3594 (2001).
Rackham, O. & Chin, J.W. A network of orthogonal ribosome·mRNA pairs . Nat. Chem. Biol. 1, 159–166 (2005).
Sleight, S.C. & Sauro, H.M. Visualization of evolutionary stability dynamics and competitive fitness of Escherichia coli engineered with randomized multigene circuits . ACS Synth. Biol. 2, 519–528 (2013).
Lovett, S.T., Hurley, R.L., Sutera, V.A., Aubuchon, R.H. & Lebedeva, M.A. Crossing over between regions of limited homology in Escherichia coli: RecA-dependent and RecA-independent pathways . Genetics 160, 851–859 (2002).
Sleight, S.C., Bartley, B.A., Lieviant, J.A. & Sauro, H.M. Designing and engineering evolutionary robust genetic circuits . J. Biol. Eng. 4, 12 (2010).
Arkin, A.P. & Fletcher, D.A. Fast, cheap and somewhat in control . Genome Biol. 7, 114 (2006).
Dong, H., Nilsson, L. & Kurland, C.G. Gratuitous overexpression of genes in Escherichia coli leads to growth inhibition and ribosome destruction . J. Bacteriol. 177, 1497–1504 (1995).
Mather, W.H., Hasty, J., Tsimring, L.S. & Williams, R.J. Translational cross talk in gene networks . Biophys. J. 104, 2564–2572 (2013).
Grigorova, I.L., Phleger, N.J., Mutalik, V.K. & Gross, C.A. Insights into transcriptional regulation and σ competition from an equilibrium model of RNA polymerase binding to DNA . Proc. Natl. Acad. Sci. USA 103, 5332–5337 (2006).
Levine, J.H., Lin, Y. & Elowitz, M.B. Functional roles of pulsing in genetic circuits . Science 342, 1193–1200 (2013).
Jayanthi, S., Nilgiriwala, K.S. & Del Vecchio, D. Retroactivity controls the temporal dynamics of gene transcription . ACS Synth. Biol. 2, 431–441 (2013).
Cardinale, S., Joachimiak, M.P. & Arkin, A.P. Effects of genetic variation on the E. coli host-circuit interface . Cell Rep. 4, 231–237 (2013).
Canton, B., Labno, A. & Endy, D. Refinement and standardization of synthetic biological parts and devices . Nat. Biotechnol. 26, 787–793 (2008).
Tagami, H., Inada, T., Kunimura, T. & Aiba, H. Glucose lowers CRP* levels resulting in repression of the lac operon in cells lacking cAMP . Mol. Microbiol. 17, 251–258 (1995).
Purcell, O., Grierson, C.S., di Bernardo, M. & Savery, N.J. Temperature dependence of ssrA-tag mediated protein degradation . J. Biol. Eng. 6, 10 (2012).
Kelly, J.R. et al. Measuring the activity of BioBrick promoters using an in vivo reference standard . J. Biol. Eng. 3, 4 (2009).
Cho, B.-K., Charusanti, P., Herrgård, M.J. & Palsson, B.Ø. Microbial regulatory and metabolic networks . Curr. Opin. Biotechnol. 18, 360–364 (2007).
Yaman, F., Bhatia, S., Adler, A., Densmore, D. & Beal, J. Automated selection of synthetic biology parts for genetic regulatory networks . ACS Synth. Biol. 1, 332–344 (2012).
Bhatia, S. & Densmore, D. Pigeon: a design visualizer for synthetic biology . ACS Synth. Biol. 2, 348–350 (2013).
Huynh, L., Tsoukalas, A., Köppe, M. & Tagkopoulos, I. SBROME: a scalable optimization and module matching framework for automated biosystems design . ACS Synth. Biol. 2, 263–273 (2013).
Roehner, N. & Myers, C.J. A methodology to annotate systems biology markup language models with the synthetic biology open language . ACS Synth. Biol. 3, 57–66 (2014).
Arkin, A.P. A wise consistency: engineering biology for conformity, reliability, predictability . Curr. Opin. Chem. Biol. 17, 893–901 (2013).
Ceroni, F., Furini, S., Stefan, A., Hochkoeppler, A. & Giordano, E. A synthetic post-transcriptional controller to explore the modular design of gene circuits . ACS Synth. Biol. 1, 163–171 (2012).
Seo, J.-H. et al. Multiple-omic data analysis of Klebsiella pneumoniae MGH 78578 reveals its transcriptional architecture and regulatory features . BMC Genomics 13, 679 (2012).
Ingolia, N.T., Brar, G.A., Rouskin, S., McGeachy, A.M. & Weissman, J.S. The ribosome profiling strategy for monitoring translation in vivo by deep sequencing of ribosome-protected mRNA fragments . Nat. Protoc. 7, 1534–1550 (2012).
Becker, A.H., Oh, E., Weissman, J.S., Kramer, G. & Bukau, B. Selective ribosome profiling as a tool for studying the interaction of chaperones and targeting factors with nascent polypeptide chains and ribosomes . Nat. Protoc. 8, 2212–2239 (2013).
Siegal-Gaskins, D., Noireaux, V. & Murray, R.M. Biomolecular resource utilization in elementary cell-free gene circuits . in Proc. Am. Control Conf. 1531–1536 (IEEE, 2013).
Karzbrun, E., Shin, J., Bar-Ziv, R.H. & Noireaux, V. Coarse-grained dynamics of protein synthesis in a cell-free system . Phys. Rev. Lett. 106, 048104 (2011).
Noireaux, V., Bar-Ziv, R. & Libchaber, A. Principles of cell-free genetic circuit assembly . Proc. Natl. Acad. Sci. USA 100, 12672–12677 (2003).
Karig, D.K., Jung, S.-Y., Srijanto, B., Collier, C.P. & Simpson, M.L. Probing cell-free gene expression noise in femtoliter volumes . ACS Synth. Biol. 2, 497–505 (2013).
Lentini, R. et al. Fluorescent proteins and in vitro genetic organization for cell-free synthetic biology . ACS Synth. Biol. 2, 482–489 (2013).
Niederholtmeyer, H., Xu, L. & Maerkl, S.J. Real-time mRNA measurement during an in vitro transcription and translation reaction using binary probes . ACS Synth. Biol. 2, 411–417 (2013).
Chizzolini, F., Forlin, M., Cecchi, D. & Mansy, S.S. Gene position more strongly influences cell-free protein expression from operons than T7 transcriptional promoter strength . ACS Synth. Biol. 10.1021/sb4000977 (27 November 2013).
Sun, Z.Z., Yeung, E., Hayes, C.A., Noireaux, V. & Murray, R.M. Linear DNA for rapid prototyping of synthetic biological circuits in an Escherichia coli based TX-TL cell-free system . ACS Synth. Biol. 10.1021/sb400131a (22 November 2013).
Peralta-Yahya, P.P. et al. Identification and microbial production of a terpene-based advanced biofuel . Nat. Commun. 2, 483 (2011).
Jonnalagadda, S.B., Becker, J.U., Sel'kov, E.E. & Betz, A. Flux regulation in glycogen-induced oscillatory glycolysis in cell-free extracts of Saccharomyces carlsbergensis . Biosystems 15, 49–58 (1982).
Loo, L.W.M. et al. cis-Expression QTL analysis of established colorectal cancer risk variants in colon tumors and adjacent normal tissue . PLoS ONE 7, e30477 (2012).
Klavins, E. Proportional-integral control of stochastic gene regulatory networks . in Proc. IEEE Conf. Decis. Control 2547–2553 (IEEE, 2010).
Bernard, P. & Couturier, M. Cell killing by the F plasmid CcdB protein involves poisoning of DNA-topoisomerase II complexes . J. Mol. Biol. 226, 735–745 (1992).
Hussein, R. & Lim, H.N. Direct comparison of small RNA and transcription factor signaling . Nucleic Acids Res. 40, 7269–7279 (2012).
Hucka, M. et al. The systems biology markup language (SBML): a medium for representation and exchange of biochemical network models . Bioinformatics 19, 524–531 (2003).
Gottesman, S., Roche, E., Zhou, Y. & Sauer, R.T. The ClpXP and ClpAP proteases degrade proteins with carboxy-terminal peptide tails added by the SsrA-tagging system . Genes Dev. 12, 1338–1347 (1998).
Tabor, J.J., Bayer, T.S., Simpson, Z.B., Levy, M. & Ellington, A.D. Engineering stochasticity in gene expression . Mol. Biosys. 4, 754–761 (2008).
Scott, M., Gunderson, C.W., Mateescu, E.M., Zhang, Z. & Hwa, T. Interdependence of cell growth and gene expression: origins and consequences . Science 330, 1099–1102 (2010).
C.A.V. and J.A.N.B. are supported by the US National Institute of General Medical Sciences (NIGMS grant P50 GMO98792 and R01 GM095765), Office of Naval Research (ONR) Multidisciplinary University Research Initiative (MURI grant 4500000552) and US National Science Foundation (NSF) Synthetic Biology Engineering Research Center (SynBERC EEC0540879) and by Life Technologies (A114510). J.A.N.B. is supported by an NSF Graduate Research Fellowship.
Synthetic Biology Center, Department of Biological Engineering, Massachusetts Institute of Technology, Cambridge, Massachusetts, USA.
Correspondence to Christopher A Voigt.
Models used to generate figure 3 – the model is in SBML format and can be opened and run with SBML software.
Models used to generate figures 3 and 4 – the model is in SBML format and can be opened and run with SBML software.

References: V. 
 V. 
 V. 
 V. 
 V. 
 V. 
 V.