SynBioFromLeukipposInstitute

*A collection of some synbio papers*

*Is There Anything Unique in the Ethics of Synthetic Biology?*D Heyd - Perspectives in Biology and Medicine, 2012Abstract Synthetic biology does not create any ethical dilemmas that have not already been raised in the development of practices such as genetic screening, genetic engineering, and other interventions in the evolutionary processes. The issue is, nevertheless, ethically ...
http://bit.ly/12ZtwKo *Synthetic Biology*: *A Challenge to Mechanical Explanations in Biology?*M Morange - Perspectives in Biology and Medicine, 2012Abstract In their plans to modify organisms, synthetic biologists have contrasted engineering and tinkering. By drawing this contrast between their endeavors and what has happened during the evolution of organisms by natural selection, they underline the novelty of their ...
http://bit.ly/ZycMUw *Synthetic Biology*: *A Jewish View*S Glick - Perspectives in Biology and Medicine, 2012Abstract The discipline of synthetic biology may be one of the most dramatic advances of the past few decades. It represents a radical upgrading of humankind's ability to manipulate the world in which we live. The potential for benefits to society is enormous, but the risks for ...
http://bit.ly/WvjKL2 *Synthetic Biology and the Golem of Prague*: *Philosophical Reflections on a Suggestive Metaphor*U Charpa - Perspectives in Biology and Medicine, 2012Abstract Not only the public debate about science but even the way scientists conceive their own work is to some extent determined by cultural images. In the case of synthetic biology, literary figures like the Golem of Prague and its successors, such as Frankenstein's ...http://bit.ly/Ygkmor

*Ethical and Regulatory Challenges Posed by Synthetic Biology*B Rager-Zisman - Perspectives in Biology and Medicine, 2012Abstract Synthetic biology is a relatively new science with tremendous potential to change how we view and know the life sciences, but like many developing technologies, it has provoked ethical concerns from the scientific community and the public and confronts ...
http://bit.ly/XQHdap

picture source:

Cell picture show Synthetic Biologyhttp://www.cell.com/cell_picture_show-synthbio

by
Gregory Linshiz, Alex Goldberg, Tania Konry, Nathan J. Hillson

"Synthetic biology is a nascent field that emerged in earnest only around the turn of the millennium. It aims to engineer new biological systems and impart new biological functionality, often through genetic modifications. The design and construction of new biological systems is a complex, multistep process, requiring multidisciplinary collaborative efforts from "fusion" scientists who have formal training in computer science or engineering, as well as hands-on biological expertise. The public has high expectations for synthetic biology and eagerly anticipates the development of solutions to the major challenges facing humanity. This article discusses laboratory practices and the conduct of research in synthetic biology. It argues that the fusion science approach, which integrates biology with computer science and engineering best practices, including standardization, process optimization, computer-aided design and laboratory automation, miniaturization, and systematic management, will increase the predictability and reproducibility of experiments and lead to breakthroughs in the construction of new biological systems. The article also discusses several successful fusion projects, including the development of software tools for DNA construction design automation, recursive DNA construction, and the development of integrated microfluidics systems."

http://bit.ly/ZyaDbD

by
Cheryl P. Goldbeck †, Heather M. Jensen , Michaela A. TerAvest , Nicole Beedle, Yancey Appling †, Matt Hepler, Guillaume Cambray , Vivek Mutalik, Largus T. Angenent , and Caroline M. Ajo-Franklin

"Introduction of the electron transfer complex MtrCAB from Shewanella oneidensis MR-1 into a heterologous host provides a modular and molecularly defined route for electrons to be transferred to an extracellular inorganic solid. However, an Escherichia coli strain expressing this pathway displayed limited control of MtrCAB expression and impaired cell growth. To overcome these limitations and to improve heterologous extracellular electron transfer, we used an E. coli host with a more tunable induction system and a panel of constitutive promoters to generate a library of strains that separately transcribe the mtr and cytochrome c maturation (ccm) operons over 3 orders of magnitude. From this library, we identified strains that show 2.2 times higher levels of MtrC and MtrA and that have improved cell growth. We find that a 300-fold decrease in the efficiency of MtrC and MtrA synthesis with increasing mtr promoter activity critically limits the maximum expression level of MtrC and MtrA. We also tested the extracellular electron transfer capabilities of a subset of the strains using a three-electrode microbial electrochemical system. Interestingly, the strain with improved cell growth and fewer morphological changes generated the largest maximal current per cfu, rather than the strain with more MtrC and MtrA. This strain also showed 30-fold greater maximal current per cfu than its ccm-only control strain. Thus, the conditions for optimal MtrCAB expression and anode reduction are distinct, and minimal perturbations to cell morphology are correlated with improved extracellular electron transfer in E. coli."

http://bit.ly/10S4Bn4

by
Ewen Callaway

"The latest shopping website is open for business, offering unusual wares: DNA tools to help biologists to engineer life.

 The DNA sequences — which allow precise control of gene activity in the bacterium Escherichia coli — are the first output of BIOFAB, based in Emery­ville, California, which calls itself “the world’s first biological design-build facility”. Launched in 2009 with a US$1.4-million grant from the US National Science Foundation, BIOFAB aims to advance synthetic biology by creating standard biological ‘parts’ in the form of DNA sequences that control gene expression. These standard sequences should allow biologists to engineer cells that can make medicines and perform other useful tasks simply by plugging in various sets of genes. The sequences are meant to overcome a key barrier to synthetic biology: genes inserted into an organism do not behave predictably, even in such a well-understood workhorse as E. coli. “You would think after a generation of genetic engineering, expressing genes with precision in an organism as well utilized as E. coliwould be pretty straightforward. It turns out it’s not,” says BIOFAB co-director Drew Endy, a synthetic biologist at Stanford University in California. Over the past three decades, scientists have amassed collections of these sequences and used them to express genes in which they are interested. Some sequences tend to be ‘strong’ and others ‘weak’, resulting in varying levels of RNA and protein being produced. But a team led by Endy and BIOFAB co-director Adam Arkin, of Lawrence Berkeley National Laboratory in Berkeley, California, has found that the activities of those sequences are far from predictable. In two papers published online this week in Nature Methods1, 2, the team reports inserting many different combinations of promoters and RBS sequences in front of genes encoding fluorescent proteins, and then measuring the level of protein that was made. “It was a bloody mess,” says Arkin, with each promoter–RBS combination having varying effects depending on the gene. He and Endy also cite an earlier finding that a scientist hoping to express a protein at a particular level has just a 50% chance of producing the required amount within a factor of two. Such hit-or-miss expression poses a major challenge to synthetic biologists who would like to create genetic circuits involving dozens of genes. As a solution, the BIOFAB team designed promoter and RBS sequences for E. colithat do not interfere with downstream DNA, so that their effects are independent of the specific gene they are paired with. The sequences should provide scientists with a much tighter grip on gene expression, offering around a 93% chance of hitting a desired level of expression within a factor of two2. Researchers can obtain the sequences for free online (see http://www.biofab.org/data), and Arkin says that some of his colleagues are already finding them useful. Endy and Arkin’s team also devised a statistical method1 to measure the variability in the performance of their promoter and RBS sequences, and indeed any genetic part to be used in synthetic-biology applications. The method should allow researchers to create a kind of specification sheet for each biological part, making it easier for scientists to develop and share their work. Randy Rettberg, a synthetic biologist at the Massachusetts Institute of Technology in Cambridge, who has worked with Endy on similar projects, says that more labs should follow BIOFAB’s lead and industrialize the production of biological parts. And synthetic biologist Alistair Elfick of the University of Edinburgh, UK, says that the BIOFAB products should help synthetic biologists to design bigger and more complicated circuits....."

http://bit.ly/ZHgtt6

*Library of synthetic transcriptional AND gates built with split T7 RNA polymerase mutants*

by
David L. Shisa and Matthew R. Bennett

"The construction of synthetic gene circuits relies on our ability to engineer regulatory architectures that are orthogonal to the host’s native regulatory pathways. However, as synthetic gene circuits become larger and more complicated, we are limited by the small number of parts, especially transcription factors, that work well in the context of the circuit. The current repertoire of transcription factors consists of a limited selection of activators and repressors, making the implementation of transcriptional logic a complicated and component-intensive process. To address this, we modified bacteriophage T7 RNA polymerase (T7 RNAP) to create a library of transcriptional AND gates for use in Escherichia coli by first splitting the protein and then mutating the DNA recognition domain of the C-terminal fragment to alter its promoter specificity. We first demonstrate that split T7 RNAP is active in vivo and compare it with full-length enzyme. We then create a library of mutant split T7 RNAPs that have a range of activities when used in combination with a complimentary set of altered T7-specific promoters. Finally, we assay the two-input function of both wild-type and mutant split T7 RNAPs and find that regulated expression of the N- and C-terminal fragments of the split T7 RNAPs creates AND logic in each case. This work demonstrates that mutant split T7 RNAP can be used as a transcriptional AND gate and introduces a unique library of components for use in synthetic gene circuits."

http://bit.ly/10JMweE

comment: 
*Wetware advances*: *Biological logic gate built by splitting viral gene*

by
John Timmer 

"In recent years, researchers in the messy world of biology have been able to build systems that function like the clean, binary switches on computer chips—and we've covered a number of reports in this area. Unfortunately, most of these share a significant limitation: they rely on proteins from bacteria that act as switches to turn genes on and off under specific conditions. We know about only a limited number of these genetic switches, which may set a severe limit on the number of logical operations we can string together inside a cell.

 A paper in this week's PNAS describes a system that may allow us to get around this limitation. The new method takes a protein from a virus that infects bacteria and cuts it in two, making a pair of genes (A and B) that each produce part of the mature protein. The two parts then act as a biological version of an AND logic gate, with output (in the form of protein activity) present only when both A and B interact. When either or both A and B are missing, the output is off. In biological terms, the inputs usually involve a simple molecule that can be sensed by proteins inside a bacteria. This paper, for example, used two kinds of sugars (arabinose and lactose). When the sugars are present, they attach to proteins inside the cell, activating genes that are controlled by those proteins. To make an AND gate, you need to design a bit of biology that can respond to both of these signals—it should be active only when both a gene regulated by arabinose and a gene regulated by lactose are each active. This has been done in a variety of ways in the past, but the authors of the new paper (both faculty at Rice University) come up with a clever scheme for doing so. It's clever in part because it's so remarkably simple. The uses of a T7 bacteriophage The new work relies on a gene from a virus called the T7 bacteriophage that infects bacteria. Instead of relying on host proteins to ensure that its genes are turned into RNA (and thus into proteins), T7 carries its own gene for a protein that transcribes DNA into RNA. This gene, called (wait for it) T7 RNA polymerase, recognizes DNA sequences on the virus, sticks to them, and then starts copying the DNA nearby into RNA. If you move the sequences it recognizes somewhere else—into the bacterial genome, onto a construct you supply—it will start copying there instead. T7 RNA polymerase therefore makes a good output. When it's active, you can use it to turn on a variety of genes, thus coordinating a significant response to your logic. But how do you get T7 RNA polymerase to respond to the two different inputs required for an AND gate? You break it in two. Scientists who were studying T7 RNA polymerase had found that, during purification, it would sometimes get cut into two different pieces, one about four times the size of the other. Either of the parts on its own is inactive, but if you put them together they stick, and the resulting aggregate is active (that is, it would bind to the appropriate DNA sequences and start making RNA, albeit at a slightly slower rate than the intact protein). It turns out this also works if the two parts are encoded by completely separate genes. You can encode the larger T7 RNA polymerase fragment in a gene that responds to arabinose and the smaller fragment in a gene that responds to lactose. A functional T7 RNA polymerase will only be present when both sugars are present, so you've made your biological AND gate. By putting a fluorescent protein under the control of T7 RNA polymerase, the authors were able to show that this worked as expected. The cells glowed green only when both sugars were added. That on its own is pretty good, although not so much better than some previous work in the field. The great part of this system is its flexibility. Because T7 RNA polymerase has been studied extensively, researchers have identified a variety of mutations that alter the protein's ability to bond to specific DNA sequences. A single change in the right location can thus switch T7 RNA polymerase from sticking to (for example) a sequence that includes GACG to one that includes the sequence GCAT. Other changes in the T7 RNA polymerase can alter the sequence it recognizes even further. Instead of relying on different proteins for every logical operation you need to do (which will quickly exhaust your supply of tractable proteins), you can now build up logic using different forms of T7 RNA polymerase, each recognizing a somewhat different sequence. This doesn't necessarily help with alternate logic operations, like NOT, but having a larger array of potential tools can only make designing biological circuitry easier."

http://ars.to/YqqEhc

Fig http://bit.ly/16q2tqB

by
Jacob Beal, Ron Weiss, Douglas Densmore, Aaron Adler, Evan Appleton, Jonathan Babb, Swapnil Bhatia, Noah Davidsohn, Traci Haddock, Joseph Loyall, Richard Schantz, Viktor Vasilev, and Fusun Yaman

"We present a workflow for the design and production of biological networks from high-level program specifications. The workflow is based on a sequence of intermediate models that incrementally translate high-level specifications into DNA samples that implement them. We identify algorithms for translating between adjacent models and implement them as a set of software tools, organized into a four-stage toolchain: Specification, Compilation, Part Assignment, and Assembly. The specification stage begins with a Boolean logic computation specified in the Proto programming language. The compilation stage uses a library of network motifs and cellular platforms, also specified in Proto, to transform the program into an optimized Abstract Genetic Regulatory Network (AGRN) that implements the programmed behavior. The part assignment stage assigns DNA parts to the AGRN, drawing the parts from a database for the target cellular platform, to create a DNA sequence implementing the AGRN. Finally, the assembly stage computes an optimized assembly plan to create the DNA sequence from available part samples, yielding a protocol for producing a sample of engineered plasmids with robotics assistance. Our workflow is the first to automate the production of biological networks from a high-level program specification. Furthermore, the workflow’s modular design allows the same program to be realized on different cellular platforms simply by swapping workflow configurations. We validated our workflow by specifying a small-molecule sensor-reporter program and verifying the resulting plasmids in both HEK 293 mammalian cells and in E. coli bacterial cells."

http://bit.ly/13ZTAV6

by
Roberta Lentini , Michele Forlin , Laura Martini , Cristina Del Bianco , Amy C. Spencer , Domenica Torino , and Sheref S. Mansy 

"To facilitate the construction of cell-free genetic devices, we evaluated the ability of 17 different fluorescent proteins to give easily detectable fluorescence signals in real-time from in vitro transcription-translation reactions with a minimal system consisting of T7 RNA polymerase and E. coli translation machinery, i.e., the PUREsystem. The data were used to construct a ratiometric fluorescence assay to quantify the effect of genetic organization on in vitro expression levels. Synthetic operons with varied spacing and sequence composition between two genes that coded for fluorescent proteins were then assembled. The resulting data indicated which restriction sites and where the restriction sites should be placed in order to build genetic devices in a manner that does not interfere with protein expression. Other simple design rules were identified, such as the spacing and sequence composition influences of regions upstream and downstream of ribosome binding sites and the ability of non-AUG start codons to function in vitro."

http://bit.ly/12DfUo5

by
Kevin V. Solomon, Tae Seok Moon, Brian Ma, Tarielle M. Sanders, and Kristala L. J. Prather 

"Tuning expression of competing endogenous pathways has been identified as an effective strategy in the optimization of heterologous production pathways. However, intervention at the first step of glycolysis, where no alternate routes of carbon utilization exist, remains unexplored. In this work we have engineered a viable E. coli host that decouples glucose transport and phosphorylation, enabling independent control of glucose flux to a heterologous pathway of interest through glucokinase (glk) expression. Using community sourced and curated promoters, glk expression was varied over a 3-fold range while maintaining cellular viability. The effects of glk expression on the productivity of a model glucose-consuming pathway were also studied. Through control of glycolytic flux we were able to explore a number of cellular phenotypes and vary the yield of our model pathway by up to 2-fold in a controllable manner."

http://bit.ly/WlDEcJ

by
Thomas C. Williams , Lars K. Nielsen , and Claudia E. Vickers 

"Population-density-dependent control of gene expression, or quorum sensing, is widespread in nature and is used to coordinate complex population-wide phenotypes through space and time. We have engineered quorum sensing in S. cerevisiae by rewiring the native pheromone communication system that is normally used by haploid cells to detect potential mating partners. In our system, populations consisting of only mating type “a” cells produce and respond to extracellular α-type pheromone by arresting growth and expressing GFP in a population-density-dependent manner. Positive feedback quorum sensing dynamics were tuned by varying α-pheromone production levels using different versions of the pheromone-responsive FUS1 promoter as well as different versions of pheromone genes (mfα1 or mfα2). In a second system, pheromone communication was rendered conditional upon the presence of aromatic amino acids in the growth medium by controlling α-pheromone expression with the aromatic amino acid responsive ARO9 promoter. In these circuits, pheromone communication and response could be fine-tuned according to aromatic amino acid type and concentration. The genetic control programs developed here are responsive to dynamic spatiotemporal and chemical cellular environments, resulting in up-regulation of gene expression. These programs could be used to control biochemical pathways for the production of fuels and chemicals that are toxic or place a heavy metabolic burden on cell growth."

http://bit.ly/ZNRbY8

by
M. Mitchell Waldrop

"When campus president Wallace Loh walked into Juan Uriagereka's office last August, he got right to the point. “We need courses for this thing — yesterday!”

 Uriagereka, associate provost for faculty affairs at the University of Maryland in College Park, knew exactly what his boss meant. Campus administrators around the world had been buzzing for months about massive open online courses, or MOOCs: Internet-based teaching programmes designed to handle thousands of students simultaneously, in part using the tactics of social-networking websites. To supplement video lectures, much of the learning comes from online comments, questions and discussions. Participants even mark one another's tests.MOOCs had exploded into the academic consciousness in summer 2011, when a free artificial-intelligence course offered by Stanford University in California attracted 160,000 students from around the world — 23,000 of whom finished it. Now, Coursera in Mountain View, California — one of the three researcher-led start-up companies actively developing MOOCs — was inviting the University of Maryland to submit up to five courses for broadcast on its software platform. Loh wanted in. “He was very clear,” says Uriagereka. “We needed to be a part of this...."

http://bit.ly/12TXxva

by
Tovar JD.

"Peptide self-assembly is a powerful method to create functional nanoscale materials such as optoelectronically relevant organic nanostructures. The enormous potential that may come from bringing π-conjugated electronic function into biological environments is poised to impact cell and tissue engineering, biosensors, and related biomedical applications. However, very little synthetic guidance is available with respect to uniting these two different materials sets in a generally applicable manner. In this Account, I describe my group's work to synthesize and assemble peptidic nanostructures built around organic electronic elements. The Account begins with a very brief background to the area of supramolecular electronics, followed by a description of areas where these nanomaterials could be useful in biology. I then discuss the synthetic approaches that we utilized to embed a variety of π-electron units directly within peptide backbones. A key supramolecular challenge with respect to subsequent self-assembly of these new molecules is balancing electrostatic contributions within the resulting nanomaterials, because the suitable geometries for stabilizing peptide assemblies may not necessarily correspond to those suitable for maximizing intermolecular π-electron interactions. Regardless of the respective magnitudes of these two major influences, the assembly paradigm is fairly robust. Variation of the π-electron units and the peptide sequences that make up the "peptide-π-peptide" triblock molecules consistently leads to fairly uniform tape-like nanostructures that maintain strong electronic coupling among the component π-electron units. We explored a diverse range of π-electron units spanning fluorescent oligo(phenylene vinylene)s, electron-accepting rylene diimides, and hole-transporting oligothiophenes. I then describe the characterization of the nanomaterials that form after molecular self-assembly in order to understand their internal structures, electronic interactions, and morphologies as existing within self-supporting hydrogel matrices. I also describe how a facile shearing process provided globally aligned macroscopic collections of one-dimensional electronic fibrils in hydrogel matrices. These general assembly processes influence intermolecular π-stacking among the embedded chromophores, and the assemblies themselves can facilitate the covalent cross-linking and polymerization (for example, of reactive diyne units). The latter offers an exciting possibility to create peptidic nanostructures comprised of single polymer chains. Finally, I discuss electronic properties as manifested in the interactions of transition dipoles within the nanomaterials and electrical properties resulting from field-effect gating. The ability to tune the observable electrical properties of the nanostructures externally will allow for their transition to in vitro or in vivo platforms as a powerful new approach to regulating biological interactions at the nanoscale."

http://bit.ly/ZM1NZJ