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Synthetic Biology of Antimicrobial Discovery

Synthetic Biology of Antimicrobial Discovery | SynBioFromLeukipposInstitute | Scoop.it

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Bijan Zakeri and Timothy K. Lu

"Antibiotic discovery has a storied history. From the discovery of penicillin by Sir Alexander Fleming to the relentless quest for antibiotics by Selman Waksman, the stories have become like folklore used to inspire future generations of scientists. However, recent discovery pipelines have run dry at a time when multidrug-resistant pathogens are on the rise. Nature has proven to be a valuable reservoir of antimicrobial agents, which are primarily produced by modularized biochemical pathways. Such modularization is well suited to remodeling by an interdisciplinary approach that spans science and engineering. Herein, we discuss the biological engineering of small molecules, peptides, and non-traditional antimicrobials and provide an overview of the growing applicability of synthetic biology to antimicrobials discovery."
http://bit.ly/SGvY0g

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Genome writing project confronts technology hurdles

More than 250 scientists, entrepreneurs, and ethicists assembled in New York City last week to discuss Genome Project–write (GP-write), which aims to build large stretches of synthetic DNA and put them to work in cells. The still-unfunded project promises better and cheaper DNA-writing technology that could help treat and study diseases or create more sustainable food and energy sources. But questions remain about whether GP-write should pursue an original goal, championed by co-founder and futurist Andrew Hessel, of assembling the 3 billion DNA bases of the human genome. In the meantime, the initiative aims to support technology-advancing "pilot projects" that modify cells from various organisms. But technical challenges—from the cost of DNA synthesis to genetic circuit design—loom large.
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Genome Partitioner: A web tool for multi-level partitioning of large-scale DNA constructs for synthetic biology applications

Recent advances in lower-cost DNA synthesis techniques have enabled new innovations in the field of synthetic biology. Still, efficient design and higher-order assembly of genome-scale DNA constructs remains a labor-intensive process. Given the complexity, computer assisted design tools that fragment large DNA sequences into fabricable DNA blocks are needed to pave the way towards streamlined assembly of biological systems. Here, we present the Genome Partitioner software implemented as a web-based interface that permits multi-level partitioning of genome-scale DNA designs. Without the need for specialized computing skills, biologists can submit their DNA designs to a fully automated pipeline that generates the optimal retrosynthetic route for higher-order DNA assembly. To test the algorithm, we partitioned a 783 kb Caulobacter crescentus genome design. We validated the partitioning strategy by assembling a 20 kb test segment encompassing a difficult to synthesize DNA sequence. Successful assembly from 1 kb subblocks into the 20 kb segment highlights the effectiveness of the Genome Partitioner for reducing synthesis costs and timelines for higher-order DNA assembly. The Genome Partitioner is broadly applicable to translate DNA designs into ready to order sequences that can be assembled with standardized protocols, thus offering new opportunities to harness the diversity of microbial genomes for synthetic biology applications. The Genome Partitioner web tool can be accessed at https://christenlab.ethz.ch/GenomePartitioner.
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Microsoft Reportedly Wants to Use DNA for Cloud Data Storage

Microsoft Reportedly Wants to Use DNA for Cloud Data Storage | SynBioFromLeukipposInstitute | Scoop.it
In the not-so-distant future, next time you want to back up your work to Microsoft’s cloud, you might be storing it on a few snippets of DNA.
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Leonid Fridman's curator insight, May 23, 4:57 PM
הנה שימוש בטכנולוגיה בימינו שלנו בעדי לשצור קבצים בעתיד אנחנו נשתמש בדגימת די אן איי
 
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Synthetic biology: Return to sender

Temporal control of gene expression is critical for cellular function and fate determination. Some genes, such as the Notch effector Hes1, exhibit an oscillating pattern of gene expression, marked by rapid mRNA synthesis and degradation due to negative feedback. Optogenetic approaches have enabled the generation of artificial oscillations with rapid spatial–temporal precision, whereas the use of bioluminescent or fluorescent reporters allows detection of oscillations at the single-cell level. However, it is not clear whether this oscillatory information can be transfe…
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Cell-free synbio: a technology whose time has come | PLOS Synthetic Biology Community

Cell-free synbio: a technology whose time has come | PLOS Synthetic Biology Community | SynBioFromLeukipposInstitute | Scoop.it
‘It’s like instant noodles – just add water’ exclaims Dr. Keith Pardee, now an assistant professor at the University of Toronto. He is describing a small black object developed during his postdoc in the Collins lab. This unassuming device is one of the most advanced biosensors ever built – it is able to detect the presence of the Zika virus. This is achieved by using an RNA toehold switch which provides the molecular precision required to identify and outbreaks and help guide efforts to combat virus’ spread. But things get really interesting when you look at the transformative technology behind the sensor.

 

Rather than use the common approaches of polymerase chain reaction (PCR) or a genetically modified bacteria to detect the virus, Keith decided to use a ‘cell free’ system. The principle is simple: grow up bacteria, smash open the cells, and use the contents to perform reactions in a test tube or on paper (for a guide on how to make extracts see this JoVE article). This has a number of advantages over the alternatives: unlike PCR, the assay can be used in remote locations without the need for expensive lab equipment, thereby allowing instantaneous field testing, and unlike genetically modified bacteria, cell-free systems avoid concerns about uncontrolled escape of genetically modified organisms into the environment.
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Towards implementation of cellular automata in Microbial Fuel Cells

Towards implementation of cellular automata in Microbial Fuel Cells | SynBioFromLeukipposInstitute | Scoop.it
The Microbial Fuel Cell (MFC) is a bio-electrochemical transducer converting waste products into electricity using microbial communities. Cellular Automaton (CA) is a uniform array of finite-state machines that update their states in discrete time depending on states of their closest neighbors by the same rule. Arrays of MFCs could, in principle, act as massive-parallel computing devices with local connectivity between elementary processors. We provide a theoretical design of such a parallel processor by implementing CA in MFCs. We have chosen Conway’s Game of Life as the ‘benchmark’ CA because this is the most popular CA which also exhibits an enormously rich spectrum of patterns. Each cell of the Game of Life CA is realized using two MFCs. The MFCs are linked electrically and hydraulically. The model is verified via simulation of an electrical circuit demonstrating equivalent behaviours. The design is a first step towards future implementations of fully autonomous biological computing devices with massive parallelism. The energy independence of such devices counteracts their somewhat slow transitions—compared to silicon circuitry—between the different states during computation.
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“Addressing the ethical issues raised by synthetic human entities with embryo-like features” Human life starts day 0

The "14-day rule" for embryo research stipulates that experiments with intact human embryos must not allow them to develop beyond 14 days or the appearance of the primitive streak. However, recent experiments showing that suitably cultured human pluripotent stem cells can self-organize and recapitulate embryonic features have highlighted difficulties with the 14-day rule and led to calls for its reassessment. Here we argue that these and related experiments raise more foundational issues that cannot be fixed by adjusting the 14-day rule, because the framework underlying the rule cannot adequately describe the ways by which synthetic human entities with embryo-like features (SHEEFs) might develop morally concerning features through altered forms of development. We propose that limits on research with SHEEFs be based as directly as possible on the generation of such features, and recommend that the research and bioethics communities lead a wide-ranging inquiry aimed at mapping out solutions to the ethical problems raised by them.
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Use of processed data to design an orderly logic gate to construct plasmids in GenoCAD

Rapid developments have been made in synthetic biology within the past two decades, particularly in combination with chemistry, computer science, and other disciplines. Genetic components and internal features have been a main focus of research for synthetic biologists. Logic gates can be applied in various disciplines, but have not yet been used for plasmid design. GenoCAD is a computer-aided design software programme for synthetic biology that can be used to design complex structures. Thus, in this study, the authors analysed a large, commonly used data set containing over 70,000 feature sequences and eventually obtained comprehensive information for a complete data set without redundancy. By analysing the internal feature sequences, the authors input the most representative data in the GenoCAD platform, along with design rules and grammar for constructing high-quality practical parts. Additionally, the orderly logic gate for building biological parts designed in this study may be useful for professionals and non-professionals and may have applications in the design of a new biological computer. Finally, the authors compared the constructed plasmid with other successful examples in BLAST and PlasMapper software to demonstrate the rationality of the orderly logic gate.
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Generating CRISPR mouse models: Challenges and solutions

Generating CRISPR mouse models: Challenges and solutions | SynBioFromLeukipposInstitute | Scoop.it
The discovery of the CRISPR/Cas9 system in bacteria has ignited the world of gene editing. It has been applied to the modification of a growing range of organisms, with none found yet that can resist its powers. In particular, CRISPR technology has revolutionized the process for creating genetically modified mice—the workhorses for in vivo cancer research—allowing for shorter timelines and in many cases, more efficient procedures.
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CRISPR enzymes act like Pac-Man to chew up RNA

CRISPR enzymes act like Pac-Man to chew up RNA | SynBioFromLeukipposInstitute | Scoop.it
Once activated, 10 newly described CRISPR enzymes behave like Pac-Man to run amok and destroy RNA. The findings could be useful for detecting infectious viruses.

The new enzymes are variants of a CRISPR protein, Cas13a. In September, scientists reported in the journal Nature that Cas13a could be used to detect specific sequences of RNA, such as those from a virus. They showed that once it binds to its target RNA, it begins to indiscriminately cut up all RNA linked to a reporter molecule, making it fluoresce to allow signal detection.

Researchers subsequently paired CRISPR-Cas13a with RNA amplification, and showed that the system—which they dubbed Sherlock—could see viral RNA at extremely low concentrations to detect the presence of dengue and Zika, for example.

Such a system could be used to detect any type of RNA, including RNA distinctive of cancer cells, researchers say.
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The Gordon Research Seminar on Synthetic Biology

The Gordon Research Seminar on Synthetic Biology | SynBioFromLeukipposInstitute | Scoop.it
The Gordon Research Seminar on Synthetic Biology is a unique forum for graduate students, post-docs, and other scientists with comparable levels of experience and education to present and exchange new data and cutting edge ideas.
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Design of synthetic epigenetic circuits featuring memory effects and reversible switching based on DNA methylation

Design of synthetic epigenetic circuits featuring memory effects and reversible switching based on DNA methylation | SynBioFromLeukipposInstitute | Scoop.it
Recording systems would allow synthetic organisms to store a ‘memory’ of a past event for future reference.
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Cell-free technology startup founded by former SynBio SRI member awarded funding by RebelBio. — Synthetic Biology in Cambridge

Cell-free technology startup founded by former SynBio SRI member awarded funding by RebelBio. — Synthetic Biology in Cambridge | SynBioFromLeukipposInstitute | Scoop.it
Cell-Free Tech is a brand new start up company specialising in giving people the ability to do biological research, without the need for expensive tools and infrastructure. Based at the Microbiology Department of the University College Cork, Cell-free Tech is part of RebelBio, an accelerator programme that helps life sciences innovators, academics, biomakers and citizen scientists to change the world with biology.
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Base‐modified nucleic acids as a powerful tool for synthetic biology and biotechnology

The ability of various nucleoside triphosphate analogs of deoxyguanosine and deoxycytidine with 7-deazadeoxyadenosine (A1) and 5-chlorodeoxyuridine (T1) to serve as substrates for Taq DNA polymerase was evaluated. The triphosphate set constituted of A1, T1, and 7-deazadeoxyguanosine with either 5-methyldeoxy-cytidine or 5-fluorodeoxycytidine was successfully employed in the polymerase chain reaction (PCR) of 1.5 kb fragments as well as random oligonucleotide libraries. Another effective combination of triphosphates for the synthesis of 1 kb PCR product was A1, T1, deoxyinosine, and 5-bromodeoxycytidine. In vivo experiments using an antibiotic-resistant gene containing the latter set demonstrated that the bacterial machinery accepts fully modified sequences as genetic templates. Moreover, the ability of the base-modified segments to selectively protect DNA from cleavage by restriction endonucleases was shown. This evidence can be used to regulate the endonuclease cleavage pattern.
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Lab-grown blood stem cells produced at last

Lab-grown blood stem cells produced at last | SynBioFromLeukipposInstitute | Scoop.it
After 20 years of trying, scientists have transformed mature cells into primordial blood cells that regenerate themselves and the components of blood. The work, described today in Nature1, 2, offers hope to people with leukaemia and other blood disorders who need bone-marrow transplants but can’t find a compatible donor. If the findings translate into the clinic, these patients could receive lab-grown versions of their own healthy cells.

One team, led by stem-cell biologist George Daley of Boston Children’s Hospital in Massachusetts, created human cells that act like blood stem cells, although they are not identical to those found in nature1. A second team, led by stem-cell biologist Shahin Rafii of Weill Cornell Medical College in New York City, turned mature cells from mice into fully fledged blood stem cells2.

“For many years, people have figured out parts of this recipe, but they’ve never quite gotten there,” says Mick Bhatia, a stem-cell researcher at McMaster University in Hamilton, Canada, who was not involved with either study. “This is the first time researchers have checked all the boxes and made blood stem cells.”

Daley’s team chose skin cells and other cells taken from adults as their starting material. Using a standard method, they reprogrammed the cells into induced pluripotent stem (iPS) cells, which are capable of producing many other cell types. Until now, however, iPS cells have not been morphed into cells that create blood.

The next step was the novel one: Daley and his colleagues inserted seven transcription factors — genes that control other genes — into the genomes of the iPS cells. Then they injected these modified human cells into mice to develop. Twelve weeks later, the iPS cells had transformed into progenitor cells capable of making the range of cells found in human blood, including immune cells. The progenitor cells are “tantalizingly close” to naturally occurring ‘haemopoetic’ blood stem cells, says Daley.

Bhatia agrees. “It’s pretty convincing that George has figured out how to cook up human haemopoetic stem cells,” he says. “That is the holy grail.”

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Bacteria can form living materials with liquid crystals, says a new research

Bacteria can form living materials with liquid crystals, says a new research | SynBioFromLeukipposInstitute | Scoop.it
https://t.co/HZNV2j0Sdm
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Interlocked DNA topologies for nanotechnology

Interlocked DNA topologies for nanotechnology | SynBioFromLeukipposInstitute | Scoop.it
Interlocked molecular architectures are well known in supramolecular chemistry and are widely used for various applications like sensors, molecular machines and logic gates. The use of DNA for constructing these interlocked structures has increased significantly within the current decade. Because of Watson–Crick base pairing rules, DNA is an excellent material for the self-assembly of well-defined interlocked nanoarchitectures. These DNA nanostructures exhibit sufficient stability, good solubility in aqueous media, biocompatibility, and can be easily combined with other biomolecules in bio-hybrid nano-assemblies. Therefore, the study of novel DNA-based interlocked systems is of interest for nanotechnology, synthetic biology, supramolecular chemistry, biotechnology, and for sensing purposes. Here we summarize recent developments and applications of interlocked supramolecular architectures made of DNA. Examples illustrating that these systems can be precisely controlled by switching on and off the molecular motion of its mechanically trapped components are discussed. Introducing different triggers into such systems creates molecular assemblies capable of performing logic gate operations and/or catalytic activity control. Interlocked DNA-based nanostructures thus represent promising frameworks for building increasingly complex and dynamic nanomachines with highly controllable functionality.
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Biohacking & the Rise of Digital Biology

Biohacking & the Rise of Digital Biology | SynBioFromLeukipposInstitute | Scoop.it
Biohacking and digital biology are taking off with huge promise of positively impacting a wide variety of industries.
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Biobrick chain recommendations for genetic circuit design

Synthetic biology databases have collected numerous biobricks to accelerate genetic circuit design. However, selecting biobricks is a tough task. Here, we leverage the fact that these manually designed circuits can provide underlying knowledge to support biobrick selection. We propose to design a recommendation system based on the analysis of available genetic circuits, which can narrow down the biobrick selection range and provide candidate biobricks for users to choose. A recommendation strategy based on a Markov model is established to tackle this issue. Furthermore, a biobrick chain recommendation algorithm Sira is proposed that applies a dynamic programming process on a layered state transition graph to obtain the top k recommendation results. In addition, a weighted filtering strategy, WFSira, is proposed to augment the performance of Sira. The experimental results on the Registry of Standard Biological Parts show that Sira outperforms other algorithms significantly for biobrick recommendations, with approximately 30% improvement in terms of recall rate. It is also able to make biobrick chain recommendations. WFSira can further improve the recall rate of Sira by an average of 7.5% for the top 5 recommendations.
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CRISPR pioneer Jennifer Doudna shares her outlook for the groundbreaking gene-editing tool

CRISPR pioneer Jennifer Doudna shares her outlook for the groundbreaking gene-editing tool | SynBioFromLeukipposInstitute | Scoop.it
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Building a Genome from Scratch: Lessons Learned from Sc2.0 - The Center of Excellence for Engineering Biology

Building a Genome from Scratch: Lessons Learned from Sc2.0 - The Center of Excellence for Engineering Biology | SynBioFromLeukipposInstitute | Scoop.it
If anyone understands the challenges associated with powering a eukaryotic cell with a designer genome built from scratch, it’s Leslie Mitchell, a postdoc in Jef Boeke’s lab at NYU. Mitchell has been leading experimental design and technological development for the Synthetic Yeast Genome Project (Sc2.0) since 2012. In this role, she collaborates with Sc2.0 International Consortium team members spanning 4 continents to provide remote mentorship and solve challenges associated with synthetic chromosome design features and assembly.

The goal of Sc2.0 is to design, build, assemble, and test the function of an entirely synthetic designer yeast genome. Earlier this month, the Sc2.0 International Consortium announced the completed design of the Sc2.0 genome, construction of five new synthetic chromosomes, and described the 3D organization of synthetic chromosomes in the nucleus, all of which earned them the March 10 cover story for Science. To date, 30% of the Sc2.0 genome has been constructed in cells, and the team hopes to have the entire genome – all 16 chromosomes – completed before the end of 2017.

According to Mitchell, the genome engineering effort of Sc2.0 can offer some powerful lessons for GP-write, both from an organizational and technical perspective.

Organizational Insights
A well defined plan should be in place at the launch of the project, which all participants must agree upon, including funding, space, personnel, QA/standards, material transfers, publication policy, intellectual property, software, ownership of the project, training and education, and compliance with local laws. Additionally, because GP-write will involve global participation similar to Sc2.0, it will be important to maintain a collaborative and inclusive culture across international borders.

One organizational feature that has driven success of Sc2.0 is the distributive nature of the project. While Sc2.0 chromosome design is centralized, involving a close collaboration between yeast geneticists of Jef Boeke’s lab and computational biologists from Joel Bader’s lab at Johns Hopkins University, synthesis and assembly are parallelized between teams around the world. Not only does this partition the workload associated with chromosome assembly, it also distributes the financial burden as each team is responsible for obtaining their own funding.

Technical Insights
Following a common design standard has been a key strategy for Sc2.0. All design features that are written into Sc2.0 chromosomes adhere to a set of overarching design principles, the goals of which are to direct growth to wild-type levels while simultaneously increasing genome flexibility and stability.

Another lesson learned from Sc2.0 is to take on a piece-by-piece assembly strategy. Such a strategy is practical in that it’s much easier to manipulate smaller segments of DNA in vitro, for example 30-60 kb segmented into 10 kb ‘chunks’. More importantly, this strategy allows the team to evaluate synthetic DNA function in cells along the way and quickly back track to any design features that negatively impact cell fitness. For the Sc2.0 project it is now clear that despite densely spaced clusters of edits (mean distance of ~400bp), the overall design is robust as few ‘bugs’, or designer changes that affect cell fitness, have been uncovered.

Sc2.0 has also highlighted limitations of gene synthesis and how these limitations may impact our ability to design and build synthetic genomes. While cost is a much-discussed issue associated with gene synthesis, a less publicized problem is that not all sequences can be easily synthesized de novo. It all comes down to sequence composition. For instance ‘low complexity DNA’ such as homopolymer runs and regions with extreme GC or AT richness can be particularly difficult to synthesize. Of course with enough time, money and effort most DNA sequences can eventually be built; de novo designed mammalian genome-scale synthesis however will require a drop in cost by orders of magnitude together with technology improvements to enable DNA synthesis without restrictions on sequence composition.

DNA delivery is another area ripe for technology development for GP-write. Yeast cells are easily transformed and incorporate DNA into their genome readily using their natural capacity for homologous recombination. Together these features have enabled an efficient Sc2.0 DNA delivery strategy encompassing segments of 30-60kb of designer DNA. The Sc2.0 strategy can applied on some level for GP-write, but there will be delivery challenges that will need to be addressed as we move into mammalian systems, in particular with respect to the delivery of increasingly large segments of designer DNA for targeted delivery.

An additional challenge that GP-write will face with respect to building artificial mammalian chromosomes is centromere engineering. Unlike budding yeast point centromeres, which are ~125 bp and easy to synthesize, mammalian centromeres are composed of megabases of highly repetitive sequences. Mitchell and her colleagues are currently trying to overcome this engineering challenge with the goal of building mammalian artificial chromosomes that will be stable over many generations of cells. Centromere engineering strategies starting from the ‘top-down’, by minimizing native chromosomes, as well as from the ‘bottom-up’, via de novo establishment of centromere function, will both be important to pursue.

With encoded features that enable genetic flexibility and increased genomic stability, Sc2.0 will be a designer genome with new capabilities. As a result, we will soon be able to ask new biological questions – about evolution, minimal genome sequences that support viability under different conditions, and the requirement of specific genomic features such as repeats and introns. Completion of Sc2.0 will represent a major stepping-stone for GP-write, which will build on the knowledge and technological advances of this project.

We invite you to join Mitchell and her colleagues at the next GP-write meeting on May 9-10th, 2017, which will be held at the New York Genome Center. Mitchell will be speaking on the topic of Genome Engineering Foundries at 11:30 am on May 10th. View the agenda.
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Simbiotics: a multi-scale integrative platform for 3D modeling of bacterial populations

Simbiotics is a spatially explicit multi-scale modeling platform for the design, simulation and analysis of bacterial populations. Systems ranging from planktonic cells and colonies, to biofilm formation and development may be modeled. Representation of biological systems in Simbiotics is flexible, user-defined processes may be in a variety of forms depending on desired model abstraction. Simbiotics provides a library of modules such as cell geometries, physical force dynamics, genetic circuits, metabolic pathways, chemical diffusion and cell interactions. Model defined processes are integrated and scheduled for parallel multi-thead and multi-CPU execution. A virtual lab provides the modeler with analysis modules and some simulated lab equipment, enabling automation of sample interaction and data collection. An extendable and modular framework allows for the platform to be updated as novel models of bacteria are developed, coupled with an intuitive user interface to allow for model definitions with minimal programming experience. Simbiotics can integrate existing standards such as SBML, and process microscopy images to initialise the 3D spatial configuration of bacteria consortia. Two case studies, used to illustrate the platform flexibility, focus on the physical properties of the biosystems modeled. These pilot case studies demonstrate Simbiotics versatility in modeling and analysis of natural systems and as a CAD tool for synthetic biology.
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How humans could evolve to survive in space

How humans could evolve to survive in space | SynBioFromLeukipposInstitute | Scoop.it
If we hope to one day leave Earth and explore the universe, our bodies are going to have to get a lot better at surviving the harsh conditions of space. Using synthetic biology, Lisa Nip hopes to harness special powers from microbes on Earth -- such as the ability to withstand radiation -- to make humans more fit for exploring space. "We're approaching a time during which we'll have the capacity to decide our own genetic destiny," Nip says. "Augmenting the human body with new abilities is no longer a question of how, but of when."
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