scholarly journals Interfacing gene circuits with microelectronics through engineered population dynamics

2020 ◽  
Vol 6 (21) ◽  
pp. eaaz8344 ◽  
Author(s):  
M. Omar Din ◽  
Aida Martin ◽  
Ivan Razinkov ◽  
Nicholas Csicsery ◽  
Jeff Hasty
Keyword(s):  
Population Dynamics ◽  
Synthetic Biology ◽  
Integrated Circuits ◽  
Population Control ◽  
Genetic Circuits ◽  
Bacterial Colony ◽  
Bacterial Response ◽  
Impedance Profile ◽  
Changes Over Time ◽  
Engineered Bacteria

While there has been impressive progress connecting bacterial behavior with electrodes, an attractive observation to facilitate advances in synthetic biology is that the growth of a bacterial colony can be determined from impedance changes over time. Here, we interface synthetic biology with microelectronics through engineered population dynamics that regulate the accumulation of charged metabolites. We demonstrate electrical detection of the bacterial response to heavy metals via a population control circuit. We then implement this approach to a synchronized genetic oscillator where we obtain an oscillatory impedance profile from engineered bacteria. We lastly miniaturize an array of electrodes to form “bacterial integrated circuits” and demonstrate its applicability as an interface with genetic circuits. This approach paves the way for new advances in synthetic biology, analytical chemistry, and microelectronic technologies.

scholarly journals CRIMoClo plasmids for modular assembly and orthogonal chromosomal integration of synthetic circuits in Escherichia coli

2019 ◽  
Vol 13 (1) ◽  
Author(s):  
Stefano Vecchione ◽  
Georg Fritz
Keyword(s):  
Escherichia Coli ◽  
Synthetic Biology ◽  
Integrated Circuits ◽  
High Efficiency ◽  
Genetic Circuit ◽  
Dna Assembly ◽  
Chromosomal Integration ◽  
Golden Gate ◽  
Genetic Circuits ◽  
E Coli

Abstract Background Synthetic biology heavily depends on rapid and simple techniques for DNA engineering, such as Ligase Cycling Reaction (LCR), Gibson assembly and Golden Gate assembly, all of which allow for fast, multi-fragment DNA assembly. A major enhancement of Golden Gate assembly is represented by the Modular Cloning (MoClo) system that allows for simple library propagation and combinatorial construction of genetic circuits from reusable parts. Yet, one limitation of the MoClo system is that all circuits are assembled in low- and medium copy plasmids, while a rapid route to chromosomal integration is lacking. To overcome this bottleneck, here we took advantage of the conditional-replication, integration, and modular (CRIM) plasmids, which can be integrated in single copies into the chromosome of Escherichia coli and related bacteria by site-specific recombination at different phage attachment (att) sites. Results By combining the modularity of the MoClo system with the CRIM plasmids features we created a set of 32 novel CRIMoClo plasmids and benchmarked their suitability for synthetic biology applications. Using CRIMoClo plasmids we assembled and integrated a given genetic circuit into four selected phage attachment sites. Analyzing the behavior of these circuits we found essentially identical expression levels, indicating orthogonality of the loci. Using CRIMoClo plasmids and four different reporter systems, we illustrated a framework that allows for a fast and reliable sequential integration at the four selected att sites. Taking advantage of four resistance cassettes the procedure did not require recombination events between each round of integration. Finally, we assembled and genomically integrated synthetic ECF σ factor/anti-σ switches with high efficiency, showing that the growth defects observed for circuits encoded on medium-copy plasmids were alleviated. Conclusions The CRIMoClo system enables the generation of genetic circuits from reusable, MoClo-compatible parts and their integration into 4 orthogonal att sites into the genome of E. coli. Utilizing four different resistance modules the CRIMoClo system allows for easy, fast, and reliable multiple integrations. Moreover, utilizing CRIMoClo plasmids and MoClo reusable parts, we efficiently integrated and alleviated the toxicity of plasmid-borne circuits. Finally, since CRIMoClo framework allows for high flexibility, it is possible to utilize plasmid-borne and chromosomally integrated circuits simultaneously. This increases our ability to permute multiple genetic modules and allows for an easier design of complex synthetic metabolic pathways in E. coli.

scholarly journals Programming gene expression in multicellular organisms for physiology modulation through engineered bacteria

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Baizhen Gao ◽  
Qing Sun
Keyword(s):  
Gene Expression ◽  
Synthetic Biology ◽  
Logic Gates ◽  
Computational Design ◽  
Living Systems ◽  
Genetic Circuits ◽  
C Elegans ◽  
Animal Physiology ◽  
Model Animal ◽  
Engineered Bacteria

AbstractA central goal of synthetic biology is to predictably and efficiently reprogram living systems to perform computations and carry out specific biological tasks. Although there have been many advances in the bio-computational design of living systems, these advances have mainly been applied to microorganisms or cell lines; programming animal physiology remains challenging for synthetic biology because of the system complexity. Here, we present a bacteria-animal symbiont system in which engineered bacteria recognize external signals and modulate animal gene expression, twitching phenotype, and fat metabolism through RNA interference toward gfp, sbp-1, and unc-22 gene in C. elegans. By using genetic circuits in bacteria to control these RNA expressions, we are able to program the physiology of the model animal Caenorhabditis elegans with logic gates. We anticipate that engineered bacteria can be used more extensively to program animal physiology for agricultural, therapeutic, and basic science applications.

scholarly journals Programming animal physiology and behaviors through engineered bacteria

2020 ◽  
Author(s):  
Baizhen Gao ◽  
Qing Sun
Keyword(s):  
Synthetic Biology ◽  
Logic Gates ◽  
Computational Design ◽  
Living Systems ◽  
Genetic Circuits ◽  
Animal Physiology ◽  
Model Animal ◽  
And Behaviors ◽  
Animal Metabolism ◽  
Engineered Bacteria

AbstractA central goal of synthetic biology is to predictably and efficiently reprogram living systems to perform computations and carry out specific biological tasks. Although there have been many advances in the bio-computational design of living systems, these advances have mainly been applied to microorganisms or cell lines; programming animal behavior and altering animal physiology remain challenges for synthetic biology because of the system complexity. Here, we present a bacteria-animal symbiont system in which engineered bacteria recognize external signals and modulate animal gene expression, twitching behavior, and fat metabolism through RNA interference. By using genetic circuits in bacteria to control RNA expression, we programmed the physiology and behavior of the model animal Caenorhabditis elegans with logic gates. We anticipate that engineered bacteria can be used more extensively to program animal metabolism and behaviors for agricultural, therapeutic, and basic science applications.One Sentence SummaryAnimal physiology and behaviors are programmed via engineered bacteria.

scholarly journals A Synthetic Biology Approach Using Engineered Bacteria to Detect Perfluoroalkyl Substance (PFAS) Contamination in Water

2021 ◽  
Vol 186 (Supplement_1) ◽  
pp. 801-807
Author(s):  
Nathaniel A Young ◽  
Ryan L Lambert ◽  
Angela M Buch ◽  
Christen L Dahl ◽  
Jackson D Harris ◽  
...  
Keyword(s):  
Synthetic Biology ◽  
Dna Sequences ◽  
Detection System ◽  
The United States ◽  
Detection Methods ◽  
Health Concern ◽  
Contaminated Water ◽  
Engineered Bacteria ◽  
Synthetic Biology Approach ◽  
Rhodococcus Jostii

ABSTRACT Introduction Per- and polyfluoroalkyl substances (PFAS) are a class of synthetic compounds used industrially for a wide variety of applications. These PFAS compounds are very stable and persist in the environment. The PFAS contamination is a growing health issue as these compounds have been reported to impact human health and have been detected in both domestic and global water sources. Contaminated water found on military bases poses a potentially serious health concern for active duty military, their families, and the surrounding communities. Previous detection methods for PFAS in contaminated water samples require expensive and time-consuming testing protocols that limit the ability to detect this important global pollutant. The main objective of this work was to develop a novel detection system that utilizes a biological reporter and engineered bacteria as a way to rapidly and efficiently detect PFAS contamination. Materials and Methods The United States Air Force Academy International Genetically Engineered Machine team is genetically engineering Rhodococcus jostii strain RHA1 to contain novel DNA sequences composed of a propane 2-monooxygenase alpha (prmA) promoter and monomeric red fluorescent protein (mRFP). The prmA promoter is activated in the presence of PFAS and transcribes the mRFP reporter. Results The recombinant R. jostii containing the prmA promoter and mRFP reporter respond to exposure of PFAS by activating gene expression of the mRFP. At 100 µM of perfluorooctanoic acid, the mRFP expression was increased 3-fold (qRT-PCR). Rhodococcus jostii without exposure to PFAS compounds had no mRFP expression. Conclusions This novel detection system represents a synthetic biology approach to more efficiently detect PFAS in contaminated samples. With further refinement and modifications, a similar system could be readily deployed in the field around the world to detect this critical pollutant.

Population dynamics of large and small mammals: Graeme Caughley's grand vision

2009 ◽  
Vol 36 (1) ◽  
pp. 1 ◽  
Author(s):  
Charles J. Krebs
Keyword(s):  
Population Dynamics ◽  
Population Control ◽  
Population Increase ◽  
Control Mechanisms ◽  
Relative Role ◽  
Top Down ◽  
Large Herbivores ◽  
Disturbed Systems ◽  
Herbivorous Mammals ◽  
Grand Vision

Ecologists that study the population dynamics of large and small herbivorous mammals operate in two worlds that overlap only partly, and in this paper I address whether the conjecture that these worlds represent two distinct and valid paradigms is currently justified. I argue that large mammals fall into three groups depending on whether they have effective predators or not, and whether they are harvested by humans. Because of human persecution of large predators, more and more large herbivorous mammals are effectively predator-free and are controlled bottom-up by food. But in less disturbed systems, large herbivorous mammals should be controlled top-down by effective predators, and this can lead to a trophic cascade. Small herbivorous mammals have been suggested to be controlled top-down by predators but some experimental evidence has challenged this idea and replaced it with the notion that predation is one of several factors that may affect rates of population increase. Intrinsic control (territoriality, infanticide, social inhibition of breeding) appears to be common in small herbivorous mammals with altricial young but is absent in species with precocial young, in ecosystems with strong stochastic weather variation (deserts) and in areas of human-induced habitat fragmentation in agricultural monocultures. The extrinsic control of large herbivores with precocial young validates part of Graeme Caughley’s Grand Vision, but the relative role of intrinsic and extrinsic mechanisms for small herbivores with altricial young is still controversial. An improved knowledge of population control mechanisms for large and small herbivores is essential for natural resource management.

scholarly journals Properties of alternative microbial hosts used in synthetic biology: towards the design of a modular chassis

2016 ◽  
Vol 60 (4) ◽  
pp. 303-313 ◽  
Author(s):  
Juhyun Kim ◽  
Manuel Salvador ◽  
Elizabeth Saunders ◽  
Jaime González ◽  
Claudio Avignone-Rossa ◽  
...  
Keyword(s):  
Synthetic Biology ◽  
Genetic Information ◽  
Essential Element ◽  
Model Organisms ◽  
Biotechnological Applications ◽  
Cell Models ◽  
Genetic Circuits ◽  
Metabolic Diversity ◽  
Translational Machinery ◽  
Metabolic Fluxes

The chassis is the cellular host used as a recipient of engineered biological systems in synthetic biology. They are required to propagate the genetic information and to express the genes encoded in it. Despite being an essential element for the appropriate function of genetic circuits, the chassis is rarely considered in their design phase. Consequently, the circuits are transferred to model organisms commonly used in the laboratory, such as Escherichia coli, that may be suboptimal for a required function. In this review, we discuss some of the properties desirable in a versatile chassis and summarize some examples of alternative hosts for synthetic biology amenable for engineering. These properties include a suitable life style, a robust cell wall, good knowledge of its regulatory network as well as of the interplay of the host components with the exogenous circuits, and the possibility of developing whole-cell models and tuneable metabolic fluxes that could allow a better distribution of cellular resources (metabolites, ATP, nucleotides, amino acids, transcriptional and translational machinery). We highlight Pseudomonas putida, widely used in many different biotechnological applications as a prominent organism for synthetic biology due to its metabolic diversity, robustness and ease of manipulation.

scholarly journals Rock-paper-scissors: Engineered population dynamics increase genetic stability

Science ◽  
2019 ◽  
Vol 365 (6457) ◽  
pp. 1045-1049 ◽  
Author(s):  
Michael J. Liao ◽  
M. Omar Din ◽  
Lev Tsimring ◽  
Jeff Hasty
Keyword(s):  
Population Control ◽  
Single Cells ◽  
Fixation Time ◽  
Ecological Interactions ◽  
Gene Circuits ◽  
Selective Pressures ◽  
Complementary Approach ◽  
Common Strategy ◽  
The Stability ◽  
Engineered Bacteria

Advances in synthetic biology have led to an arsenal of proof-of-principle bacterial circuits that can be leveraged for applications ranging from therapeutics to bioproduction. A unifying challenge for most applications is the presence of selective pressures that lead to high mutation rates for engineered bacteria. A common strategy is to develop cloning technologies aimed at increasing the fixation time for deleterious mutations in single cells. We adopt a complementary approach that is guided by ecological interactions, whereby cyclical population control is engineered to stabilize the functionality of intracellular gene circuits. Three strains of Escherichia coli were designed such that each strain could kill or be killed by one of the other two strains. The resulting “rock-paper-scissors” dynamic demonstrates rapid cycling of strains in microfluidic devices and leads to an increase in the stability of gene circuit functionality in cell culture.

scholarly journals A Microbial Screening in Silico Method for the Fitness Step Evaluation in Evolutionary Algorithms

2020 ◽  
Vol 10 (11) ◽  
pp. 3936
Author(s):  
A. Gargantilla Becerra ◽  
R. Lahoz-Beltra
Keyword(s):  
Synthetic Biology ◽  
In Silico ◽  
Optimization Problems ◽  
Evaluation Function ◽  
Laboratory Methods ◽  
Bacterial Colony ◽  
Fitness Value ◽  
General Rules ◽  
A Cell ◽  
Microbial Screening

One of the most delicate stages of an evolutionary algorithm is the evaluation of the goodness of the solutions by some procedure providing a fitness value. However, although there are general rules, it is not always easy to find an appropriate evaluation function for a given problem. In the biological realm, today, there is a variety of experimental methods under the name of microbial screening to identify and select bacteria from their traits, as well as to obtain their fitness. In this paper, we show how given an optimization problem, a colony of synthetic bacteria or bacterial agents is able to evaluate the fitness of candidate solutions by building an evaluation function. The evaluation function is obtained simulating, in silico, a bacterial colony conducting the laboratory methods used in microbiology, biotechnology and synthetic biology to measure microbial fitness. Once the evaluation function is built, it is included in the code of the genetic algorithm as part of the fitness routine. The practical use of this approach is illustrated in two classic optimization problems. In silico routines have been programmed in Gro, a cell programming language oriented to synthetic biology, and can easily be customized to many other optimization problems.

Multimaterial bioprinting—minus the printer: Synthetic bacterial patterning with UV-responsive genetic circuits

2020 ◽  
pp. 147807712096337
Author(s):  
Gizem Gumuskaya
Keyword(s):  
Industrial Revolution ◽  
Vibrio Fischeri ◽  
Raw Material ◽  
Genetic Circuit ◽  
Elastic Tissue ◽  
Genetic Circuits ◽  
Bacterial Colony ◽  
Synthetic Dna ◽  
Macro Scale ◽  
Bacterial Lawn

In this paper, we argue that synthetic biology can help us employ living systems’ unique capacity for self-construction and biomaterial production toward developing novel architectural fabrication paradigms, in which both the raw material production and its refinement into a target structure can be merged into a single computational process embedded in the living structure itself. To demonstrate, here we introduce bioPheme, a novel biofabrication method for engineering bacteria to build biomaterial(s) of designer’s choice into arbitrary 2D geometries specified via transient UV tracing. To this end, we present the design, construction, and testing of the enabling synthetic DNA circuit, which, once inserted into a bacterial colony, allows the bacteria to execute spatial computation by interacting with one another based on the if-then rules encoded in this circuit. At the heart of this genetic circuit is a pair of UV sensor – actuator, and a pair of cell-to-cell signal transmitter – receptor modules, created with genes extracted from the virus λ Phage and marine bacterium Vibrio fischeri, respectively. These modules are wired together to help designers engineer bacteria to build macro-scale structures with seamlessly integrated biomaterials, thereby bridge the molecular and architectural scales. In this way, a bacterial lawn can be programmed to produce different objects with complementary biomaterial compositions, such as a biomineralized superstructure and an elastic tissue filling in-between. In summary, this paper focuses on how scientists’ increasing ability to harness the innate computational capacity of living cells can help designers create self-constructing structures for architectural biofabrication. Through the discussions in this paper, we aim to initiate a shift in today’s biodesign practices toward a greater appreciation and adoption of bottom-up governance of living structures. We are confident that such a paradigm shift will allow for more efficient and sustainable biofabrication systems in the 4th industrial revolution and beyond.

scholarly journals Regulating synthetic gene networks in 3D materials

2012 ◽  
Vol 109 (38) ◽  
pp. 15217-15222 ◽  
Author(s):  
Tara L. Deans ◽  
Anirudha Singh ◽  
Matthew Gibson ◽  
Jennifer H. Elisseeff
Keyword(s):  
Synthetic Biology ◽  
Gene Networks ◽  
Materials Science ◽  
Genetic Circuits ◽  
Therapeutic Applications ◽  
Regulatory Processes ◽  
Cellular Events ◽  
The Matrix

Combining synthetic biology and materials science will enable more advanced studies of cellular regulatory processes, in addition to facilitating therapeutic applications of engineered gene networks. One approach is to couple genetic inducers into biomaterials, thereby generating 3D microenvironments that are capable of controlling intrinsic and extrinsic cellular events. Here, we have engineered biomaterials to present the genetic inducer, IPTG, with different modes of activating genetic circuits in vitro and in vivo. Gene circuits were activated in materials with IPTG embedded within the scaffold walls or chemically linked to the matrix. In addition, systemic applications of IPTG were used to induce genetic circuits in cells encapsulated into materials and implanted in vivo. The flexibility of modifying biomaterials with genetic inducers allows for patterned placement of these inducers that can be used to generate distinct patterns of gene expression. Together, these genetically interactive materials can be used to characterize genetic circuits in environments that more closely mimic cells’ natural 3D settings, to better explore complex cell–matrix and cell–cell interactions, and to facilitate therapeutic applications of synthetic biology.

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