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Rationally designed families of orthogonal RNA... [Nat Chem Biol. 2012] - PubMed - NCBI

The next-generation synthetic biology applications:

Rationally designed families of orthogonal RNA regulators of translation
by
Mutalik VK, Qi L, Guimaraes JC, Lucks JB, Arkin AP.

"Our ability to routinely engineer genetic networks for applications is limited by the scarcity of highly specific and non-cross-reacting (orthogonal) gene regulators with predictable behavior. Though antisense RNAs are attractive contenders for this purpose, quantitative understanding of their specificity and sequence-function relationship sufficient for their design has been limited. Here, we use rationally designed variants of the RNA-IN-RNA-OUT antisense RNA-mediated translation system from the insertion sequence IS10 to quantify >500 RNA-RNA interactions in Escherichia coli and integrate the data set with sequence-activity modeling to identify the thermodynamic stability of the duplex and the seed region as the key determinants of specificity. Applying this model, we predict the performance of an additional ∼2,600 antisense-regulator pairs, forecast the possibility of large families of orthogonal mutants, and forward engineer and experimentally validate two RNA pairs orthogonal to an existing group of five from the training data set. We discuss the potential use of these regulators in next-generation synthetic biology applications."

http://1.usa.gov/HfPSbj

Additional page:
Duplex/RNAin/RNAout
http://bit.ly/GSQu5S

#syntheticbiology #synbio #syntheticbio #rna

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Scientists want to replace lab workhorse E. coli with the world’s fastest-growing bacterium

Scientists want to replace lab workhorse E. coli with the world’s fastest-growing bacterium | SynBioFromLeukipposInstitute | Scoop.it
<i>Vibrio natriegens</i> could save researchers valuable time
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DNA design helps create novel nanomaterials - Materials Today

DNA design helps create novel nanomaterials - Materials Today | SynBioFromLeukipposInstitute | Scoop.it
A cube, an octahedron and a prism are among the polyhedral structures, or frames, made of DNA that scientists at the US Department of Energy's (DOE) Brookhaven National Laboratory have designed to connect nanoparticles into a variety of precisely structured three-dimensional (3D) lattices. The scientists have also developed a method to integrate nanoparticles and DNA frames into interconnecting modules, expanding the diversity of possible structures.

These achievements, described in papers in Nature Materials and Nature Chemistry, could lead to the rational design of nanomaterials with enhanced or combined optical, electric and magnetic properties.

"We are aiming to create self-assembled nanostructures from blueprints," said physicist Oleg Gang, who led this research at the Center for Functional Nanomaterials (CFN), a DOE Office of Science User Facility at Brookhaven. "The structure of our nanoparticle assemblies is mostly controlled by the shape and binding properties of precisely designed DNA frames, not by the nanoparticles themselves. By enabling us to engineer different lattices and architectures without having to manipulate the particles, our method opens up great opportunities for designing nanomaterials with properties that can be enhanced by precisely organizing functional components. For example, we could create targeted light-absorbing materials that harness solar energy, or magnetic materials that increase information-storage capacity."

Gang's team has previously exploited DNA's complementary base pairing – the highly specific binding of chemical bases known by the letters A, T, G and C that make up the rungs of the DNA double-helix ‘ladder’ – to bring particles together in a precise way. Particles coated with single strands of DNA with a defined sequence of bases link to particles coated with strands with a complementary sequence (A binds with T and G binds with C) while repelling particles coated with non-complementary strands.

They have also designed 3D DNA frames whose corners have single-stranded DNA tethers to which nanoparticles coated with complementary strands can bind. When the scientists mix these nanoparticles and frames, the components self-assemble into lattices that are mainly defined by the shape of the designed frame. The Nature Materials paper describes the most recent structures achieved using this strategy.

"In our approach, we use DNA frames to promote the directional interactions between nanoparticles such that the particles connect into specific configurations that achieve the desired 3D arrays," said Ye Tian, lead author of the Nature Materials paper and a member of Gang's research team. "The geometry of each particle-linking frame is directly related to the lattice type, though the exact nature of this relationship is still being explored."

So far, the team has designed five polyhedral frame shapes – a cube, an octahedron, an elongated square bipyramid, a prism and a triangular bipyramid – but a variety of other shapes could be created.

"The idea is to construct different 3D structures (buildings) from the same nanoparticle (brick)," explained Gang. "Usually, the particles need to be modified to produce the desired structures. Our approach significantly reduces the structure's dependence on the nature of the particle, which can be gold, silver, iron, or any other inorganic material."

To design the frames, the team used DNA origami, a self-assembly technique in which short synthetic strands of DNA (staple strands) are mixed with a longer single strand of biologically-derived DNA (scaffold strand). When the scientists heat and cool this mixture, the staple strands selectively bind with or ‘staple’ the scaffold strand, causing the scaffold strand to repeatedly fold over onto itself. Computer software helps them determine the specific sequence required to ensure the DNA folds into desired shapes.

The folding of the single-stranded DNA scaffold exposes anchoring points that contain free ‘sticky’ ends – unpaired strings of DNA bases – where nanoparticles coated with complementary single-strand tethers can attach. These sticky ends can be positioned anywhere on the DNA frame, but Gang's team chose the corners so that multiple frames could be connected.

For each frame shape, the number of DNA strands linking a frame corner to an individual nanoparticle is equivalent to the number of edges converging at that corner. The cube and prism frames have three strands at each corner, for example. By producing these corner tethers with varying numbers of bases, the scientists can tune the flexibility and length of the particle-frame linkages. The interparticle distances are determined by the lengths of the frame edges, which are tens of nanometers long in the frames designed to date, but the scientists say it should be possible to tailor the frames to achieve any desired dimensions.

The scientists verified the frame structures and nanoparticle arrangements through cryo-electron microscopy (a type of microscopy conducted at very low temperatures) at the CFN and Brookhaven's Biology Department, and through x-ray scattering at the National Synchrotron Light Source II (NSLS-II), a DOE Office of Science User Facility at Brookhaven.

In the Nature Chemistry paper, Gang's team described how they used a similar DNA-based approach to create programmable two-dimensional (2D) square-like DNA frames around single nanoparticles. DNA strands inside the frames provide coupling to complementary DNA on the nanoparticles, essentially holding the particle inside the frame. Each exterior side of the frame can be individually encoded with different DNA sequences. These outer DNA strands guide frame-frame recognition and connection.

Gang likens these DNA-framed nanoparticle modules to Lego bricks. "Each module can hold a different kind of nanoparticle and interlock to other modules in different but specific ways, fully determined by the complementary pairing of the DNA bases on the sides of the frame," he said.

In other words, the frames not only determine if the nanoparticles will connect but also how they will connect. Programming the frame sides with specific DNA sequences means only frames with complementary sequences can link up.

Mixing different types of modules together can yield a variety of structures, similar to the constructs that can be generated from different Lego bricks. By creating a library of the modules, the scientists hope to be able to assemble structures on demand. The selectivity of the connections allows different types and sizes of nanoparticles to be combined into single structures.

The geometry of the connections, or how the particles are oriented in space, is very important for designing structures with desired functions. For example, optically-active nanoparticles can be arranged in a particular geometry to rotate, filter, absorb and emit light – capabilities that are relevant for applications such as display screens and solar panels.

By using different modules from their ‘library’, Gang's team has so far demonstrated the self-assembly of one-dimensional linear arrays, ‘zigzag’ chains, square-shaped and cross-shaped clusters, and 2D square lattices. The scientists have even generated a simplistic nanoscale model of Leonardo da Vinci's Vitruvian Man. "We wanted to demonstrate that complex nanoparticle architectures can be self-assembled using our approach," said Gang.

Again, the scientists used sophisticated imaging techniques – electron and atomic force microscopy at the CFN and x-ray scattering at NSLS-II – to verify that their structures were consistent with the prescribed designs and to study the assembly process in detail.

"Although many additional studies are required, our results show that we are making advances toward our goal of creating designed matter via self-assembly, including periodic particle arrays and complex nanoarchitectures with freeform shapes," said Gang. "Our approach is exciting because it is a new platform for nanoscale manufacturing, one that can lead to a variety of rationally designed functional materials."

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Synthetic biology: from mainstream to counterculture

Existing at the interface of science and engineering, synthetic biology represents a new and emerging field of mainstream biology. However, there also exists a counterculture of Do-It-Yourself biologists, citizen scientists, who have made significant inroads, particularly in the design and development of new tools and techniques. Herein, I review the development and convergence of synthetic biology's mainstream and countercultures.
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Ethical Questions Loom Over Efforts to Make a Human Genome from Scratch

Ethical Questions Loom Over Efforts to Make a Human Genome from Scratch | SynBioFromLeukipposInstitute | Scoop.it
The biggest beneficiary of a plan to fabricate a human genome from scratch could be a Massachusetts startup called Gen9 that has close ties to the authors of the still-secretive proposal.
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3 #Tech Giants Quietly Investing in Synthetic #Biology 

3 #Tech Giants Quietly Investing in Synthetic #Biology  | SynBioFromLeukipposInstitute | Scoop.it
It's time for tech investors to acknowledge the potential of sneaky R&D projects in synthetic biology at Autodesk, Intel, and Microsoft. | Limitless learning Universe
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Towards synthesis of monoterpenes and derivatives using synthetic biology

Towards synthesis of monoterpenes and derivatives using synthetic biology | SynBioFromLeukipposInstitute | Scoop.it
Synthetic biology is opening up new opportunities for the sustainable and efficient production of valuable chemicals in engineered microbial factories. Here we review the application of synthetic biology approaches to the engineering of monoterpene/monoterpenoid production, highlighting the discovery of novel catalytic building blocks, their accelerated assembly into functional pathways, general strategies for product diversification, and new methods for the optimization of productivity to economically viable levels. Together, these emerging tools allow the rapid creation of microbial production systems for a wide range of monoterpenes and their derivatives for a diversity of industrial applications.
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BioMason's bricks "grown" with sand and bacteria to hit the market next year

BioMason's bricks "grown" with sand and bacteria to hit the market next year | SynBioFromLeukipposInstitute | Scoop.it
BioMason is inching closer to commercially debuting its eco-friendly “grown” bricks, which they say could be on the market as early as next year. Since its founding in 2012, the North Carolina startup has been pushing to revolutionize the world of building materials in a way that slashes carbon emissions by using bricks grown from sand and bacteria, instead of firing traditional bricks. An increasing number of architects and builders are looking to sustainable construction materials in response to the growing demand for green buildings, and BioMason’s emission-saving biobricks fill a unique niche in the market.
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Towards synthesis of monoterpenes and derivatives using synthetic biology

Synthetic biology is opening up new opportunities for the sustainable and efficient production of valuable chemicals in engineered microbial factories. Here we review the application of synthetic biology approaches to the engineering of monoterpene/monoterpenoid production, highlighting the discovery of novel catalytic building blocks, their accelerated assembly into functional pathways, general strategies for product diversification, and new methods for the optimization of productivity to economically viable levels. Together, these emerging tools allow the rapid creation of microbial production systems for a wide range of monoterpenes and their derivatives for a diversity of industrial applications.
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Biosensor reveals multiple sources for mitochondrial NAD+

Roles of cellular nicotinamide adenine dinucleotide (NAD+) in metabolism, aging, and disease have garnered much interest, but methods have been lacking to measure the amounts of NAD+ in living cells. Cambronne et al. developed a genetically encoded biosensor that can be used to monitor concentrations of free NAD+ in various compartments of a cell (see the Perspective by Guarente). Such concentrations of NAD+ appear to be important in regulating the activity of NAD+-consuming enzymes such as sirtuins and ADP-ribosyltransferases. The authors used the sensor to demonstrate that NAD+ concentrations in mitochondria of cultured human cells can be controlled by multiple mechanisms.
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Was Darwin a Punk? A Q&A with Punker-Paleontologist Greg Graffin

Was Darwin a Punk? A Q&A with Punker-Paleontologist Greg Graffin | SynBioFromLeukipposInstitute | Scoop.it
The evolutionary biologist and lead singer for the punk rock band Bad Religion explains why there are no good songs about science and how evolution can be a guide to life
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On the Path Toward Bionic Enzymes

On the Path Toward Bionic Enzymes | SynBioFromLeukipposInstitute | Scoop.it
Berkeley Lab chemists have successfully married chemistry and biology to create reactions never before possible.
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Copyrighting DNA Is a Bad Idea

Copyrighting DNA Is a Bad Idea | SynBioFromLeukipposInstitute | Scoop.it
A few years ago, molecular biologists Jennifer Doudna and Emmanuelle Charpentier, along with a team of researchers at the University…
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Lattice engineering through nanoparticle-DNA frameworks

Lattice engineering through nanoparticle-DNA frameworks | SynBioFromLeukipposInstitute | Scoop.it
Advances in self-assembly over the past decade have demonstrated that nano- and microscale particles can be organized into a large diversity of ordered three-dimensional (3D) lattices. However, the ability to generate different desired lattice types from the same set of particles remains challenging. Here, we show that nanoparticles can be assembled into crystalline and open 3D frameworks by connecting them through designed DNA-based polyhedral frames. The geometrical shapes of the frames, combined with the DNA-assisted binding properties of their vertices, facilitate the well-defined topological connections between particles in accordance with frame geometry. With this strategy, different crystallographic lattices using the same particles can be assembled by introduction of the corresponding DNA polyhedral frames. This approach should facilitate the rational assembly of nanoscale lattices through the design of the unit cell.
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A Robotic Platform for Automated RNA Extraction and Analysis during Reporter Gene–Based Dynamic Characterization of Bacterial Promoters

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In Just 3 Days, AI Solves Biology Mystery: How Flatworms Regenerate into New Organisms

In Just 3 Days, AI Solves Biology Mystery: How Flatworms Regenerate into New Organisms | SynBioFromLeukipposInstitute | Scoop.it
A computer has solved one of biology's biggest mysteries - how a sliced up flatworm can regenerate into new organisms, and it only took it a matter of days. However, years of programming went into the tech.
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The Great Debate - What Is Life?

Richard Dawkins, J. Craig Venter, Nobel laureates Sidney Altman and Leland Hartwell, Chris McKay, Paul Davies, Lawrence Krauss, and The Science ...
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Behind Feng Zhang's research, a passion for biology

Behind Feng Zhang's research, a passion for biology | SynBioFromLeukipposInstitute | Scoop.it
A passion for biology is one of the driving forces behind the research of Chinese American synthetic biologist Feng Zhang (張鋒), one of the three recipients of the Tang Prize in Biopharmaceutical Science for the development of groundbreaking gene editing technology.

It's a passion that began in junior high school, where biology teachers inspired Zhang's enduring love for the science by telling him about the many breakthroughs in biology and biochemistry that could be of great benefit to many people.

Biology later remained at the center of his student life, whether as an undergraduate at Harvard University or as a graduate student at Stanford University, and the 34-year-old is now the youngest head of a lab at the Broad Institute in Cambridge, a high-powered genomics research center affiliated with MIT and Harvard.

"Biology is an amazing and profound system. Progress in biotechnology can improve people's health and life," Zhang said.

The CRISPR/Cas9 technology for which Zhang won the Tang Prize along with Emmanuelle Charpentier of the Max Planck Institute and Jennifer Doudna of the University of California at Berkeley, could provide some of the health breakthroughs Zhang envisions.

While the two female scientists are credited with achieving the key CRISPR breakthrough that enable researchers to edit parts of the genome, Zhang has made his mark by showing how the technology could be adapted to deal with disease by applying it to edit animal genomes and get it to work in human cells.

"CRISPR, or genome editing, is a very powerful tool," Zhang said. "We can use it to understand how genes work and how different kinds of genetic variations underlie disease."

Zhang said he is hoping that this understanding will lead to new treatments for genetic disorders, fight cancer and even develop better plants with higher yields in the long-term.

Acknowledging that gene editing technology "is still very young," the biologist said he and his team are attempting to "make it more perfect and precise" in the hope of using it to provide the "greatest benefits" to people in the future.

Zhang, who also teaches in MIT's Brain and Cognitive Sciences and Biological Engineering departments, is originally from Hebei province in China, and emigrated to the United States with his family at the age of 11 and settled in Des Moines, Iowa.

For a person like Zhang with such a fertile mind, getting an education in the U.S. had real benefits.

Talking about the National Higher Education Entrance Examination that exists in China, Zhang said he found the education system in the U.S. to be more flexible and offered more "opportunities."

"It is a very good opportunity to develop one's own interests," Zhang said.

That educational freedom and a passion for his research -- Zhang spends much of his time in the lab -- have fueled his rise in the biological engineering field, helping him earn a Tang Prize.

Zhang and the two other winners of the category will share a cash prize of NT$40 million (US$1.23 million) and a research grant of up to NT$10 million to be used within five years, and will receive medals and certificates.

The Tang Prize is only the latest of a series of honors Zhang has won.

Earlier this year, he shared the Canada Gairdner International Award with Doudna and Charpentier, often said to be a precursor to winning a Nobel prize.

In 2014, he won the National Science Foundation's Alan T. Waterman Award, the Jacob Heskel Gabbay Award in Biotechnology and Medicine (shared with Doudna and Charpentier) and the Society for Neuroscience Young Investigator Award (shared with Diana Bautista).

The biennial Tang Prize was established by Taiwanese entrepreneur Samuel Yin (尹衍樑) in 2012 to complement the Nobel Prize and to honor top researchers and leaders in four fields -- sustainable development, biopharmaceutical science, Sinology, and the rule of law.
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Engineering Therapeutic T Cells: From Synthetic Biology to Clinical Trials

Engineered T cells are currently in clinical trials to treat patients with cancer, solid organ transplants, and autoimmune diseases. However, the field is still in its infancy. The design and manufacturing of T cell therapies is not standardized and is performed mostly in academic settings by competing groups. Reliable methods to define dose and pharmacokinetics of T cell therapies need to be developed. As of mid-2016, there are no FDA approved T cell therapeutics on the market and FDA regulations are only slowly adapting to the new technologies. Further development of engineered T cell therapies requires advances in immunology, synthetic biology, manufacturing processes, and government regulation. In this review, we outline some of these challenges and discuss the contributions that pathologists can make to this emerging field.
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Hack the Planet!

Hack the Planet! | SynBioFromLeukipposInstitute | Scoop.it
Hackathons are ‘hacking’ ‘marathons’, and traditionally bring people together to try and solve a defined problem in the field of software development over a period of several days. The term first came in to use in 1999 and hackathons are now widespread, they have even been proposed as a great way for scientists to work on new ideas, form collaborations and forward their careers.
However, software problems are not the only issues to be tackled by hackathons; next week the first ‘DIYBio style’ biohackathon will convene in Cambridge, UK funded by the Cambridge Synthetic Biology Research Initiative. We will be covering the event via twitter, but to find out more I went to talk to the organiser  Thomas Meany.
SJB: Can you tell us about your background, and how you came to be involved? 
TM: I’m originally trained as a physicist and recently joined the Haseloff SynBio Lab in Cambridge. We are a highly interdisciplinary lab with a grand vision of learning how cells differentiate and make stuff. The lab is very diverse and it is not at all weird to have a background in computer science or physics. It was very encouraging to see challenges in biology that I thought I might be able to help solve. For instance, automation and reproducibility by incorporating low volume lab-on-a-chip devices has enormous potential.
However, there is no clear definition of synbio or standard route to entry apart from initiatives like the iGEM competition. This is a chance for people from all walks of life to spend a short period of time working on an interdisciplinary team to help them decide how they could fit into the synbio ecosystem.
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The Quest to Make Code Work Like Biology Just Took A Big Step

The Quest to Make Code Work Like Biology Just Took A Big Step | SynBioFromLeukipposInstitute | Scoop.it
IN THE EARLY 1970s, at Silicon Valley’s Xerox PARC, Alan Kay envisioned computer software as something akin to a biological system, a vast collection of small cells that could communicate via simple messages. Each cell would perform its own discrete task. But in communicating with the rest, it would form a more complex whole. “This is an almost foolproof way of operating,” Kay once told me. Computer programmers could build something large by focusing on something small. That’s a simpler task, and in the end, the thing you build is stronger and more efficient.

The result was a programming language called SmallTalk. Kay called it an object-oriented language—the “objects” were the cells—and it spawned so many of the languages that programmers use today, from Objective-C and Swift, which run all the apps on your Apple iPhone, to Java, Google’s language of choice on Android phones.
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RNAiFold2T: Constraint Programming design of thermo-IRES switches

RNA thermometers (RNATs) are cis-regulatory elements that change secondary structure upon temperature shift. Often involved in the regulation of heat shock, cold shock and virulence genes, RNATs constitute an interesting potential resource in synthetic biology, where engineered RNATs could prove to be useful tools in biosensors and conditional gene regulation.
RESULTS:
Solving the 2-temperature inverse folding problem is critical for RNAT engineering. Here we introduce RNAiFold2T, the first Constraint Programming (CP) and Large Neighborhood Search (LNS) algorithms to solve this problem. Benchmarking tests of RNAiFold2T against existent programs (adaptive walk and genetic algorithm) inverse folding show that our software generates two orders of magnitude more solutions, thus allowing ample exploration of the space of solutions. Subsequently, solutions can be prioritized by computing various measures, including probability of target structure in the ensemble, melting temperature, etc. Using this strategy, we rationally designed two thermosensor internal ribosome entry site (thermo-IRES) elements, whose normalized cap-independent translation efficiency is approximately 50% greater at 42 °C than 30 °C, when tested in reticulocyte lysates. Translation efficiency is lower than that of the wild-type IRES element, which on the other hand is fully resistant to temperature shift-up. This appears to be the first purely computational design of functional RNA thermoswitches, and certainly the first purely computational design of functional thermo-IRES elements.
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Designing a robust single-molecule switch

Designing a robust single-molecule switch | SynBioFromLeukipposInstitute | Scoop.it
In molecular electronics, researchers combine atomic control of molecular structure with high-precision nanofabrication techniques to connect individual molecules to tiny electrodes (1). The aim is to understand electrical transport, such as conductivity in individual molecules, and to uncover phenomena with practical utility in nanoelectronic devices. On page 1443 of this issue, Jia et al. (2) report robust, room-temperature optoelectronic switching of a single molecule connected to conducting carbon (graphene) contacts. In terms of the magnitude of the switching effect, its reversibility, and stability at room temperature, the results represent the state of the art for single-molecule electronics.
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Bristol University | BrisSynBio | 2016: Colston

Bristol University | BrisSynBio | 2016: Colston | SynBioFromLeukipposInstitute | Scoop.it
7 September, University of Bristol: Colston Research Symposium on Synthetic Biology:
https://t.co/zK3aCCMkOg
Reduced registration fees!
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CRISPR/Cas9-Based Efficient Genome Editing in Clostridium ljungdahlii, an Autotrophic Gas-Fermenting Bacterium 

CRISPR/Cas9-Based Efficient Genome Editing in Clostridium ljungdahlii, an Autotrophic… https://t.co/HDnZtZI0pi https://t.co/jJpFYmYSRM
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