FOCUS ON sYnTHETIc BIoLoGY PERSPECTIVES circuits that underpin the response of a cell TIMELINE to its environment. The ability to assemble new regulatory systems from molecular A brief history of synthetic biology components was soon envisioned5, but it was not until the molecular details of transcrip- tional regulation in bacteria were uncovered D. Ewen Cameron, Caleb J. Bashor and James J. Collins in subsequent years6 that a more concrete Abstract | The ability to rationally engineer microorganisms has been a vision, based on programmed gene long-envisioned goal dating back more than a half-century. With the genomics expression, began to take shape. Following the development of molecular revolution and rise of systems biology in the 1990s came the development of a cloning and PCR in the 1970s and 1980s, rigorous engineering discipline to create, control and programme cellular genetic manipulation became widespread behaviour. The resulting field, known as synthetic biology, has undergone dramatic in microbiology research, ostensibly offer- growth throughout the past decade and is poised to transform biotechnology and ing a technical means to engineer artificial medicine. This Timeline article charts the technological and cultural lifetime of gene regulation. However, during this pre- genomic period, research approaches that synthetic biology, with an emphasis on key breakthroughs and future challenges. were categorized as genetic engineering were mostly restricted to cloning and recom- The founding of the field of synthetic biol- strategies. In this Timeline article, we focus binant gene expression. In short, genetic ogy near the turn of the millennium was on efforts in synthetic biology that deal with engineering was not yet equipped with based on the transformational assertion that microbial systems; work in mammalian the necessary knowledge or tools to create engineering approaches — then mostly for- synthetic biology has been recently reviewed biological systems that display the diversity eign to cell and molecular biology — could elsewhere2,3. and depth of regulatory behaviour found in be used both to study cellular systems and to In this Timeline article, a brief history of microorganisms. facilitate their manipulation to productive some of the major events that have shaped By the mid‑1990s, automated DNA ends. Now more than a decade old, synthetic synthetic biology since its inception are pre- sequencing and improved computational biology has undergone considerable growth sented. We begin by describing the unique tools enabled complete microbial genomes in scope, expectation and output, and has interdisciplinary dynamics of the 1990s to be sequenced, and high-throughput tech- become a widely recognized branch of bio- that, by the end of the decade, had enticed niques for measuring RNA, protein, lipids logical research1. In many aspects, the tra- engineers from disciplines outside biology and metabolites enabled scientists to gener- jectory of the field during its first decade of to enter the wet lab and begin tinkering with ate a vast catalogue of cellular components existence has been non-linear, with periods cellular networks. We divide a chronology and their interactions. This ‘scaling‑up’ of of meaningful progress matched by epi- of the field into three distinct periods and molecular biology generated the field of sodes of inertia as design efforts have been highlight scientific and cultural milestones systems biology, as biologists and computer forced to re-orient when confronted with the for each period (FIG. 1 (TIMELINE)): first, a scientists began to combine experimentation complexity and unpredictability of engineering foundational period, in which many of the and computation to reverse-engineer cellular inside living cells. characteristic experimental and cultural networks7–9. What emerged from this enor- Although a consensus has yet to be reached features of the field were established; second, mous and continuing basic research effort on a precise definition of synthetic biology, an intermediate period, which was charac- was a view that cellular networks, although the use of molecular biology tools and tech- terized by an expansion of the field but a lag vast and intricate, were organized as a hierar- niques to forward-engineer cellular behaviour in engineering advances; and third, a recent chy of clearly discernable functional modules, has emerged as a broad identity for the era of accelerated innovation and shifting similar to many engineered systems10. field, and a set of common engineering practices, in which new technologies and Gradually, it was recognized that the approaches and laboratory practices have engineering approaches have enabled us to rational manipulation of biological systems, developed, along with a vibrant community advance towards practical applications in either by systematically tuning or rearrang- culture. Much of the foundational work in the both biotechnology and medicine. ing their modular molecular constituents, field was carried out in the model microbial could form the basis of a formal biological species Escherichia coli and Saccharomyces 1961–1999: origins of the field engineering discipline11. As a complement to cerevisiae, and these microbial systems The roots of synthetic biology can be traced the top-down approach of systems biology, a remain central in several focal areas of the to a landmark publication by Francois Jacob bottom‑up approach was envisioned, which field, including complex circuit design, and Jacques Monod in 1961 (REF. 4). Insights could draw on an ever-expanding list of metabolic engineering, minimal genome from their study of the lac operon in E. coli molecular ‘parts’ to forward-engineer regu- construction and cell-based therapeutic led them to posit the existence of regulatory latory networks. Such an approach could be NATURE REVIEWS | MICROBIOLOGY VOLUME 12 | MAY 2014 | 381 © 2014 Macmillan Publishers Limited. All rights reserved PERSPECTIVES Timeline | A brief history of synthetic biology SB1.0: the first international Earliest combinatorial synthesis conference for synthetic biology First synthetic circuits of genetic networks25 held at MIT — toggle switch and Cellular regulation by (1980s–1990s) repressilator15,16 (2002–2003) First iGEM competition held at MIT molecular networks Rise of ‘omics’ era Synthetic circuits used to study postulated by Jacob of high-throughput Autoregulatory negative- transcriptional noise during this RNA devices for modular regulation and Monod4 biology feedback circuit21 period27–29 of gene expression35 1960s 1970s 1980s 1990s 2000 2001 2002 2003 2004 2005 (1970s–1980s) Widespread use of automated First cell–cell Artemisinin Light-sensing circuit engineered in Development of DNA sequencing communication precursor pathway E. coli — bacterial photography40 molecular cloning circuit based on engineered in techniques Complete genome sequence quorum sensing30 E. coli41 Programmable ligand-controlled of S. cerevisiae117 transcript regulation by RNA36 Complete genome sequence Circuits capable of multicellular of E. coli118 pattern formation are generated38 Key to coloured boxes: technical or cultural milestones (black); circuit engineering (red); synthetic biology in metabolic engineering (green); therapeutic applications (blue); whole genome engineering (purple). E. coli, Escherichia coli; iGEM, International Genetically Engineered Machine; MAGE, multiplex automated genome engineering; MIT, Massachusetts Institute of Technology; SB1.0, Synthetic Biology 1.0; S. cerevisiae, Saccharomyces cerevisiae. used both to study the functional organiza- states in response to external signals. In regulators to combinatorially assemble tion of natural systems and to create artificial another example, Elowitz and Leibler engi- genetic circuits that display diverse logic gate regulatory networks that have potential bio- neered an oscillatory circuit that consisted of behaviour25. Seminal work by Weiss and technology and health applications12. By the a triple negative-feedback loop of sequential colleagues established methods for engineer- end of the 1990s, a small group of engineers, repressor–promoter pairs16 (FIG. 2b). Activa- ing transcription-based logic gates and did physicists and computer scientists recog- tion of the circuit, termed the repressilator, much to formalize the language and practice nized the opportunity and began to migrate resulted in the ordered, periodic oscillation of circuit engineering26. Simple circuits that into molecular biology to try their hand at of repressor protein expression. explored the relationship between gene the bench. Both the toggle and repressilator were expression and molecular noise in both constructed from a similar set of parts prokaryotic and eukaryotic genes provided 2000–2003: the foundational years (for example, inducible promoter systems) an early glimpse into the role that synthetic A convenient starting point for early syn- and used GFP expression as an output to systems could have in clarifying and expanding thetic biologists was the creation of simple monitor circuit behaviour. Model-based our understanding of basic biology27–29. gene regulatory circuits that carry out func- design was used in each case, but agreement Although mostly focused on circuit tions in an analogous manner to electrical between the model and the experimental engineering, efforts during this early period circuits13,14. The dynamics of these simple output was reached only after ‘tuning’ the began to push