scholarly journals Cell-free genetic devices confer autonomic and adaptive properties to hydrogels

2019 ◽  
Author(s):  
Colette J. Whitfield ◽  
Alice M. Banks ◽  
Gema Dura ◽  
John Love ◽  
Jonathan E. Fieldsend ◽  
...  
Keyword(s):  
Gene Expression ◽  
Protein Synthesis ◽  
Information Processing ◽  
Synthetic Biology ◽  
Smart Materials ◽  
Living Organism ◽  
Chemical Diversity ◽  
Biological Information ◽  
Self Healing ◽  
Wide Range

AbstractSmart materials are able to alter one or more of their properties in response to defined stimuli. Our ability to design and create such materials, however, does not match the diversity and specificity of responses seen within the biological domain. We propose that relocation of molecular phenomena from living cells into hydrogels can be used to confer smart functionality to materials. We establish that cell-free protein synthesis can be conducted in agarose hydrogels, that gene expression occurs throughout the material and that co-expression of genes is possible. We demonstrate that gene expression can be controlled transcriptionally (using in gel gene interactions) and translationally in response to small molecule and nucleic acid triggers. We use this system to design and build a genetic device that can alter the structural property of its chassis material in response to exogenous stimuli. Importantly, we establish that a wide range of hydrogels are appropriate chassis for cell-free synthetic biology, meaning a designer may alter both the genetic and hydrogel components according to the requirements of a given application. We probe the relationship between the physical structure of the gel and in gel protein synthesis and reveal that the material itself may act as a macromolecular crowder enhancing protein synthesis. Given the extensive range of genetically encoded information processing networks in the living kingdom and the structural and chemical diversity of hydrogels, this work establishes a model by which cell-free synthetic biology can be used to create autonomic and adaptive materials.Significance statementSmart materials have the ability to change one or more of their properties (e.g. structure, shape or function) in response to specific triggers. They have applications ranging from light-sensitive sunglasses and drug delivery systems to shape-memory alloys and self-healing coatings. The ability to programme such materials, however, is basic compared to the ability of a living organism to observe, understand and respond to its environment. Here we demonstrate the relocation of biological information processing systems from cells to materials. We achieved this by operating small, programmable genetic devices outside the confines of a living cell and inside hydrogel matrices. These results establish a method for developing materials functionally enhanced with molecular machinery from biological systems.

scholarly journals The Cognitive Lens: a primer on conceptual tools for analysing information processing in developmental and regenerative morphogenesis

2019 ◽  
Vol 374 (1774) ◽  
pp. 20180369 ◽  
Author(s):  
Santosh Manicka ◽  
Michael Levin
Keyword(s):  
Information Processing ◽  
Evolutionary Biology ◽  
Biological Information ◽  
Neural Cells ◽  
Cognitive Architectures ◽  
Least Action ◽  
Mechanistic Investigation ◽  
Wide Range ◽  
Level Information ◽  
Scale Integration

Brains exhibit plasticity, multi-scale integration of information, computation and memory, having evolved by specialization of non-neural cells that already possessed many of the same molecular components and functions. The emerging field of basal cognition provides many examples of decision-making throughout a wide range of non-neural systems. How can biological information processing across scales of size and complexity be quantitatively characterized and exploited in biomedical settings? We use pattern regulation as a context in which to introduce the Cognitive Lens—a strategy using well-established concepts from cognitive and computer science to complement mechanistic investigation in biology. To facilitate the assimilation and application of these approaches across biology, we review tools from various quantitative disciplines, including dynamical systems, information theory and least-action principles. We propose that these tools can be extended beyond neural settings to predict and control systems-level outcomes, and to understand biological patterning as a form of primitive cognition. We hypothesize that a cognitive-level information-processing view of the functions of living systems can complement reductive perspectives, improving efficient top-down control of organism-level outcomes. Exploration of the deep parallels across diverse quantitative paradigms will drive integrative advances in evolutionary biology, regenerative medicine, synthetic bioengineering, cognitive neuroscience and artificial intelligence. This article is part of the theme issue ‘Liquid brains, solid brains: How distributed cognitive architectures process information’.

scholarly journals Insuperable problems of the genetic code initially emerging in an RNA World

2017 ◽  
Author(s):  
Peter R Wills ◽  
Charles W Carter
Keyword(s):  
Protein Synthesis ◽  
Information Processing ◽  
Natural Selection ◽  
Information Transfer ◽  
Rna World ◽  
Biological Information ◽  
Universal System ◽  
Biological Information Processing ◽  
Relationship Of ◽  
The Relationship

AbstractDifferential equations for error-prone information transfer (template replication, transcription or translation) are developed in order to consider, within the theory of autocatalysis, the advent of coded protein synthesis. Variations of these equations furnish a basis for comparing the plausibility of contrasting scenarios for the emergence of tRNA aminoacylation, ultimately by enzymes, and the relationship of this process with the origin of the universal system of molecular biological information processing embodied in the Central Dogma. The hypothetical RNA World does not furnish an adequate basis for explaining how this system came into being, but principles of self-organisation that transcend Darwinian natural selection furnish an unexpectedly robust basis for a rapid, concerted transition to genetic coding from a peptide•RNA world.

scholarly journals Dual UTR-A novel 5′ untranslated region design for synthetic biology applications

2020 ◽  
Vol 5 (1) ◽  
Author(s):  
Simone Balzer Le ◽  
Ingerid Onsager ◽  
Jon Andreas Lorentzen ◽  
Rahmi Lale
Keyword(s):  
Gene Expression ◽  
Synthetic Biology ◽  
De Novo ◽  
Expression Profiles ◽  
Regulation Of Gene Expression ◽  
Design Concept ◽  
Fine Tuning ◽  
The Novel ◽  
Wide Range ◽  
Application Potential

Abstract Bacterial 5′ untranslated regions of mRNA (UTR) involve in a complex regulation of gene expression; however, the exact sequence features contributing to gene regulation are not yet fully understood. In this study, we report the design of a novel 5′ UTR, dual UTR, utilizing the transcriptional and translational characteristics of 5′ UTRs in a single expression cassette. The dual UTR consists of two 5′ UTRs, each separately leading to either increase in transcription or translation of the reporter, that are separated by a spacer region, enabling de novo translation initiation. We rationally create dual UTRs with a wide range of expression profiles and demonstrate the functionality of the novel design concept in Escherichia coli and Pseudomonas putida using different promoter systems and coding sequences. Overall, we demonstrate the application potential of dual UTR design concept in various synthetic biology applications ranging from fine-tuning of gene expression to maximization of protein production.

scholarly journals Positive-feedback, ratiometric biosensor expression improves high-throughput metabolite-producer screening efficiency in yeast

2017 ◽  
Vol 2 (1) ◽  
Author(s):  
Thomas C. Williams ◽  
Xin Xu ◽  
Martin Ostrowski ◽  
Isak S. Pretorius ◽  
Ian T. Paulsen
Keyword(s):  
Gene Expression ◽  
Synthetic Biology ◽  
High Throughput ◽  
Positive Feedback ◽  
High Throughput Screening ◽  
Transcriptional Regulator ◽  
Gfp Expression ◽  
Dynamic Regulation ◽  
Wide Range ◽  
Screening Efficiency

Biosensors are valuable and versatile tools in synthetic biology that are used to modulate gene expression in response to a wide range of stimuli. Ligand responsive transcription factors are a class of biosensor that can be used to couple intracellular metabolite concentration with gene expression to enable dynamic regulation and high-throughput metabolite producer screening. We have established the Saccharomyces cerevisiae WAR1 transcriptional regulator and PDR12 promoter as an organic acid biosensor that can be used to detect varying levels of para-hydroxybenzoic acid (PHBA) production from the shikimate pathway and output green fluorescent protein (GFP) expression in response. The dynamic range of GFP expression in response to PHBA was dramatically increased by engineering positive-feedback expression of the WAR1 transcriptional regulator from its target PDR12 promoter. In addition, the noise in GFP expression at the population-level was controlled by normalising GFP fluorescence to constitutively expressed mCherry fluorescence within each cell. These biosensor modifications increased the high-throughput screening efficiency of yeast cells engineered to produce PHBA by 5,000-fold, enabling accurate fluorescence activated cell sorting isolation of producer cells that were mixed at a ratio of 1 in 10,000 with non-producers. Positive-feedback, ratiometric transcriptional regulator expression is likely applicable to many other transcription-factor/promoter pairs used in synthetic biology and metabolic engineering for both dynamic regulation and high-throughput screening applications.

scholarly journals Robust gene expression control in human cells with a novel universal TetR aptamer splicing module

2019 ◽  
Vol 47 (20) ◽  
pp. e132-e132 ◽  
Author(s):  
Adam A Mol ◽  
Florian Groher ◽  
Britta Schreiber ◽  
Ciaran Rühmkorff ◽  
Beatrix Suess
Keyword(s):  
Gene Expression ◽  
Synthetic Biology ◽  
Dynamic Range ◽  
Intron Retention ◽  
Human Cells ◽  
Fine Tuning ◽  
Genomic Context ◽  
Mammalian Cell Lines ◽  
Gene Expression Control ◽  
Wide Range

Abstract Fine-tuning of gene expression is desirable for a wide range of applications in synthetic biology. In this context, RNA regulatory devices provide a powerful and highly functional tool. We developed a versatile, robust and reversible device to control gene expression by splicing regulation in human cells using an aptamer that is recognized by the Tet repressor TetR. Upon insertion in proximity to the 5′ splice site, intron retention can be controlled via the binding of TetR to the aptamer. Although we were able to demonstrate regulation for different introns, the genomic context had a major impact on regulation. In consequence, we advanced the aptamer to develop a splice device. Our novel device contains the aptamer integrated into a context of exonic and intronic sequences that create and maintain an environment allowing a reliable and robust splicing event. The exon-born, additional amino acids will then be cleaved off by a self-cleaving peptide. This design allows portability of the splicing device, which we confirmed by demonstrating its functionality in different gene contexts. Intriguingly, our splicing device shows a high dynamic range and low basal activity, i.e. desirable features that often prove a major challenge when implementing synthetic biology in mammalian cell lines.

scholarly journals Synthetic Biology Bicistronic Designs Support Gene Expression Equally Well in vitro and in vivo

2020 ◽  
Vol 17 (1) ◽  
pp. 13-20
Author(s):  
Owen Koucky ◽  
Jacob Wagner ◽  
Sofia Aguilera ◽  
Benjamin Bashaw ◽  
Queena Chen ◽  
...  
Keyword(s):  
Gene Expression ◽  
Protein Synthesis ◽  
Synthetic Biology ◽  
Protein Production ◽  
Persistent Problem ◽  
Free Protein ◽  
Microfluidic Droplets ◽  
Cell Free Protein Synthesis

Synthetic biology integrates molecular biology tools and an engineering mindset to address challenges in medicine, agriculture, bioremediation, and biomanufacturing. A persistent problem in synthetic biology has been designing genetic circuits that produce predictable levels of protein. In 2013, Mutalik and colleagues developed bicistronic designs (BCDs) that make protein production more predicable in bacterial cells (in vivo). With the growing interest in producing proteins outside of cells (in vitro), we wanted to know if BCDs would work as predictably in cell-free protein synthesis (CFPS) as they do in E. coli cells. We tested 20 BCDs in CFPS and found they performed very similarly in vitro and in vivo. As a step toward developing methods for protein production in artificial cells, we also tested 3 BCDs inside nanoliter-scaled microfluidic droplets. The BCDs worked well in the microfluidic droplets, but their relative protein production levels were not as predictable as expected. These results suggest that the conditions under which gene expression happens in droplets result in a different relationship between genetic control elements such as BCDs and protein production than exists in batch CFPS or in cells. KEYWORDS: Bicistronic Design; Synthetic Biology; Cell-Free Protein Synthesis; Microfluidics

scholarly journals Cognitive and emotional information processing: protein synthesis and gene expression

2007 ◽  
Vol 584 (2) ◽  
pp. 389-400 ◽  
Author(s):  
Sreedharan Sajikumar ◽  
Sheeja Navakkode ◽  
Volker Korz ◽  
Julietta U. Frey
Keyword(s):  
Gene Expression ◽  
Protein Synthesis ◽  
Information Processing ◽  
Emotional Information ◽  
Emotional Information Processing

EXTRACTION OF CO-BEHAVING GENES BY SIMILARITY ENSEMBLES

2017 ◽  
Vol 25 (03) ◽  
pp. 479-494
Author(s):  
MOSLEM MOHAMMADI-JENGHARA ◽  
HOSSEIN EBRAHIMPOUR-KOMLEH
Keyword(s):  
Gene Expression ◽  
Candidate Genes ◽  
Biological Information ◽  
Data Sets ◽  
Expression Data ◽  
Yeast Gene ◽  
Ensemble Technique ◽  
Wide Range ◽  
The Impact

Microarray technology is used as a source of data for a wide range of biology studies. Useful biological information can be extracted from the analysis of microarray data, namely, the impact of a particular gene expression on the expression of other genes or the determination of expressed genes under different conditions. The purpose of this paper is to find co-behavioral genes in different data sets for different times and conditions. In other words, genes with similar behavior, same increase or decrease, under different medical, stress, and time conditions in terms of expression are determined. Multi-valued discretization of expression data was used for extracting genes with identical behavior. The algorithm proposed in this study is based on data and methods ensemble. The data ensemble technique was used to extract candidate genes with identical behavior. Other methods were also applied on all the data sets; as a result, many co-behavioral candidate genes with different similarity and correlation values were identified. Finally, the ultimate output was created from the ensemble of different methods. By applying the algorithm on yeast gene expression data, meaningful relations among genes were extracted.

scholarly journals Dual UTR-A novel 5′ untranslated region design for synthetic biology applications

2019 ◽  
Author(s):  
Simone Balzer Le ◽  
Ingerid Onsager ◽  
Jon Andreas Lorentzen ◽  
Rahmi Lale
Keyword(s):  
Gene Expression ◽  
Synthetic Biology ◽  
De Novo ◽  
Expression Profiles ◽  
Regulation Of Gene Expression ◽  
Design Concept ◽  
Fine Tuning ◽  
The Novel ◽  
Wide Range ◽  
Application Potential

ABSTRACTBacterial 5′ untranslated regions of mRNA (UTR) involve in a complex regulation of gene expression; however, the exact sequence features contributing to gene regulation are not yet fully understood. In this study, we report the design of a novel 5′ UTR, dual UTR, utilising the transcriptional and translational characteristics of 5′ UTRs in a single expression cassette. The dual UTR consists of two 5′ UTRs, each separately leading to either increase in transcription or translation of the reporter, that are separated by a spacer region, enabling de novo translation initiation. We rationally create dual UTRs with a wide range of expression profiles and demonstrate the functionality of the novel design concept in Escherichia coli and in Pseudomonas putida using different promoter systems and coding sequences. Overall, we demonstrate the application potential of dual UTR design concept in various synthetic biology applications ranging from fine-tuning of gene expression to maximisation of protein production.

scholarly journals Rapid and scalable preparation of bacterial lysates for cell-free gene expression

2017 ◽  
Author(s):  
Andriy Didovyk ◽  
Taishi Tonooka ◽  
Lev Tsimring ◽  
Jeff Hasty
Keyword(s):  
Gene Expression ◽  
Synthetic Biology ◽  
Cost Effective ◽  
Diverse Range ◽  
Highly Active ◽  
Freeze Dried ◽  
Wide Range ◽  
Specialized Equipment ◽  
Free Gene ◽  
Bacterial Lysates

AbstractCell-free gene expression systems are emerging as an important platform for a diverse range of synthetic biology and biotechnology applications, including production of robust field-ready biosensors. Here, we combine programmed cellular autolysis with a freeze-thaw or freeze-dry cycle to create a practical, reproducible, and a labor- and cost-effective approach for rapid production of bacterial lysates for cell-free gene expression. Using this method, ro-bust and highly active bacterial cell lysates can be produced without specialized equipment at a wide range of scales, making cell-free gene expression easily and broadly accessible. More-over, live autolysis strain can be freeze-dried directly and subsequently lysed upon rehydration to produce active lysate. We demonstrate the utility of autolysates for synthetic biology by reg-ulating protein production and degradation, implementing quorum sensing, and showing quan-titative protection of linear DNA templates by GamS protein. To allow versatile and sensitive β-galactosidase (LacZ) based readout we produce autolysates with no detectable background LacZ activity and use them to produce sensitive mercury(II) biosensors with LacZ-mediated colorimetric and fluorescent outputs. The autolysis approach can facilitate wider adoption of cell-free technology for cell-free gene expression as well as other synthetic biology and biotechnology applications, such as metabolic engineering, natural product biosynthesis, or proteomics.

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