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A guide to CRISPR gene activation : Wyss Institute at Harvard

A guide to CRISPR gene activation : Wyss Institute at Harvard | SynBioFromLeukipposInstitute | Scoop.it
Wyss Institute at Harvard
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Scientists Want to Bring Back Woolly Mammoths — But Why?

Scientists Want to Bring Back Woolly Mammoths — But Why? | SynBioFromLeukipposInstitute | Scoop.it
Two teams are working on "de-extinction" projects that could one day lead to another of the giant mammals walking the Earth.
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JBEI, UCSD scientists develop systems biology-based workflow to improve biofuels productivity

JBEI, UCSD scientists develop systems biology-based workflow to improve biofuels productivity | SynBioFromLeukipposInstitute | Scoop.it
Researchers at the US Department of Energy (DOE)’s Joint BioEnergy Institute (JBEI), in collaboration with researchers at the University of California, San Diego, have developed a workflow that integrates various “omics” data and genome-scale...
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Programming Surface Chemistry with Engineered Cells - ACS Synthetic Biology

We have developed synthetic gene networks that enable engineered cells to selectively program surface chemistry. E. coli were engineered to upregulate biotin synthase, and therefore biotin synthesis, upon biochemical induction. Additionally, two different functionalized surfaces were developed that utilized binding between biotin and streptavidin to regulate enzyme assembly on programmable surfaces. When combined, the interactions between engineered cells and surfaces demonstrated that synthetic biology can be used to engineer cells that selectively control and modify molecular assembly by exploiting surface chemistry. Our system is highly modular and has the potential to influence fields ranging from tissue engineering to drug development and delivery.
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Influence of Pichia pastoris cellular material on polymerase chain reaction performance as a synthetic biology standard for genome monitoring

Advances in synthetic genomics are now well underway in yeasts due to the low cost of synthetic DNA. These new capabilities also bring greater need for quantitating the presence, loss and rearrangement of loci within synthetic yeast genomes. Methods for achieving this will ideally; i) be robust to industrial settings, ii) adhere to a global standard and iii) be sufficiently rapid to enable at-line monitoring during cell growth. The methylotrophic yeast Pichia pastoris (P. pastoris) is increasingly used for industrial production of biotherapeutic proteins so we sought to answer the following questions for this particular yeast species. Is time-consuming DNA purification necessary to obtain accurate end-point polymerase chain reaction (e-pPCR) and quantitative PCR (qPCR) data? Can the novel linear regression of efficiency qPCR method (LRE qPCR), which has properties desirable in a synthetic biology standard, match the accuracy of conventional qPCR? Does cell cultivation scale influence PCR performance? To answer these questions we performed e-pPCR and qPCR in the presence and absence of cellular material disrupted by a mild 30s sonication procedure. The e-pPCR limit of detection (LOD) for a genomic target locus was 50 pg (4.91x103 copies) of purified genomic DNA (gDNA) but the presence of cellular material reduced this sensitivity sixfold to 300 pg gDNA (2.95x104 copies). LRE qPCR matched the accuracy of a conventional standard curve qPCR method. The presence of material from bioreactor cultivation of up to OD600 = 80 did not significantly compromise the accuracy of LRE qPCR. We conclude that a simple and rapid cell disruption step is sufficient to render P. pastoris samples of up to OD600 = 80 amenable to analysis using LRE qPCR which we propose as a synthetic biology standard.
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Full speed ahead to the City on the Hill

Full speed ahead to the City on the Hill | SynBioFromLeukipposInstitute | Scoop.it
With the help of synthetic biology, the city of the future can potentially be engineered less like a conglomerate of machines and more like a natural system. Cells can be engineered to sense and respond to environmental signals. Communication networks through sensors can be embedded in the city infrastructure. Protocell technology, a new material that possesses some of the properties of living systems, can be manipulated to grow architecture. We might see self-repairing architecture.
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Stanford-NIST collaboration aims to give the bio-economy a big boost by measuring tiny things

Stanford-NIST collaboration aims to give the bio-economy a big boost by measuring tiny things | SynBioFromLeukipposInstitute | Scoop.it
The Joint Initiative for Metrology in Biology, located at Stanford, will bring together academic, government and industrial scientists to improve the measurement techniques, or metrology, of molecular products and processes to facilitate advances...
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Networking biofabricated systems through molecular communication

Networking biofabricated systems through molecular communication
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Bioinformatics for the synthetic biology of natural products: integrating across the Design-Build-Test cycle

Covering: 2000 to 2016Progress in synthetic biology is enabled by powerful bioinformatics tools allowing the integration of the design, build and test stages of the biological engineering cycle. In this review we illustrate how this integration can be achieved, with a particular focus on natural products discovery and production. Bioinformatics tools for the DESIGN and BUILD stages include tools for the selection, synthesis, assembly and optimization of parts (enzymes and regulatory elements), devices (pathways) and systems (chassis). TEST tools include those for screening, identification and quantification of metabolites for rapid prototyping. The main advantages and limitations of these tools as well as their interoperability capabilities are highlighted.
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The Bioeconomy: A New Bioindustrial Revolution - CPI

The Bioeconomy: A New Bioindustrial Revolution - CPI | SynBioFromLeukipposInstitute | Scoop.it
How governments are embracing the bioindustrial revolution to grow and build their economies.
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Should we redesign humans?

Should we redesign humans? | SynBioFromLeukipposInstitute | Scoop.it
The age of bioengineering is upon us, with scientists' understanding of how to engineer cells, tissues and organs improving at a rapid pace. Here, how this could affect the future of our physical bodies.
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High-sensitivity assay for Hg (II) and Ag (I) ion detection: A new class of droplet digital PCR logic gates for an intelligent DNA calculator

The first example of droplet digital PCR logic gates (“YES”, “OR” and “AND”) for Hg (II) and Ag (I) ion detection has been constructed based on two amplification events triggered by a metal-ion-mediated base mispairing (T-Hg(II)-T and C-Ag(I)-C). In this work, Hg(II) and Ag(I) were used as the input, and the “true” hierarchical colors or “false” green were the output. Through accurate molecular recognition and high sensitivity amplification, positive droplets were generated by droplet digital PCR and viewed as the basis of hierarchical digital signals. Based on this principle, YES gate for Hg(II) (or Ag(I)) detection, OR gate for Hg(II) or Ag(I) detection and AND gate for Hg(II) and Ag(I) detection were developed, and their sensitively and selectivity were reported. The results indicate that the ddPCR logic system developed based on the different indicators for Hg(II) and Ag(I) ions provides a useful strategy for developing advanced detection methods, which are promising for multiplex metal ion analysis and intelligent DNA calculator design applications.
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AuNPs modified, disposable, ITO based biosensor: Early diagnosis of heat shock protein 70

This paper describes a novel, simple, and disposable immunosensor based on indium-tin oxide (ITO) sheets modified with gold nanoparticles to sensitively analyze heat shock protein 70 (HSP70), a potential biomarker that could be evaluated in diagnosis of some carcinomas. Disposable ITO coated Polyethylene terephthalate (PET) electrodes were used and modified with gold nanoparticles in order to construct the biosensors. Optimization and characterization steps were analyzed by electrochemical techniques such as electrochemical impedance spectroscopy (EIS) and cyclic voltammetry (CV). Surface morphology of the biosensor was also identified by electrochemical methods, scanning electron microscopy (SEM), and atomic force microscopy (AFM). To interpret binding characterization of HSP70 to anti-HSP70 single frequency impedance method was successfully operated. Moreover, the proposed HSP70 immunosensor acquired good stability, repeatability, and reproducibility. Ultimately, proposed biosensor was introduced to real human serum samples to determine HSP70 sensitively and accurately.
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Is the World Ready for Synthetic Life? Scientists Plan to Create Whole Genomes

Is the World Ready for Synthetic Life? Scientists Plan to Create Whole Genomes | SynBioFromLeukipposInstitute | Scoop.it
Last weekend, an invite-only group of about 150 experts convened privately at Harvard. Behind closed doors, they discussed the prospect of designing and building an entire human genome from scratch, using only a computer, a DNA synthesizer and raw materials.

The artificial genome would then be inserted into a living human cell to replace its natural DNA. The hope is that the cell “reboots,” changing its biological processes to operate based on instructions provided by the artificial DNA.

In other words, we may soon be looking at the first “artificial human cell.”

But the goal is not just Human 2.0. The project, “HGP-Write: Testing Large Synthetic Genomes in Cells,” also hopes to develop powerful new tools that push synthetic biology into exponential growth on an industrial scale. If successful, we won’t only have the biological tools to design humans as a species — we would have the ability to redesign the living world.

Manufacturing Life

At its core, synthetic biology is a marriage between engineering principles and biotechnology. If DNA sequencing is about reading DNA, genetic engineering is about editing DNA, synthetic biology is about programming new DNA — regardless of its original source — to build new forms of life.

Synthetic biologists view DNA and genes as standard biological bricks that can be used interchangeably to create and modify living cells.

The field has a plug-and-play mentality, says Dr. Jay Kiesling, a pioneer of synthetic engineering at the University of California at Berkeley. “When your hard drive dies, you can go to the nearest computer store, buy a new one, swap it out,” he says, “Why shouldn’t we use biological parts in the same way?”

To accelerate the field, Kiesling and colleagues are putting together a database of standardized DNA pieces — dubbed “BioBricks” — that can be used as puzzle pieces to assemble genetic material completely new to nature.

To Kiesling and others in the field, synthetic biology is like developing a new programming language. Cells are hardware, while DNA is the software that makes them run. With enough knowledge about how genes work, synthetic biologists believe that they will be able to write genetic programs from scratch, allowing them to build new organisms, alter nature and even guide the course of human evolution.

Similar to genetic engineering, synthetic biology gives scientists the power to tinker with natural DNA. The difference is mostly scale: genetic editing is a cut-and-paste process that adds foreign genes or changes the letters in existing genes. Often, only a few sites are changed.

Synthetic biology, on the other hand, creates genes from scratch. This allows scientists far more opportunities to make extensive changes to known genes, or even design their own. The possibilities are nearly endless.

Biodrugs, Biofuels, BioCrops

The explosion of synthetic biology in the past decade has already churned out results that thrilled both scientists and corporations.

Back in 2003, Kiesling published one of the earliest proof-of-concept studies demonstrating the power of the approach. He focused on a chemical called artemisinin, a powerful anti-malaria drug extracted from sweet wormwood that’s often the last line of defense against the disease.

Yet despite numerous attempts at cultivating the plant, yields remain extremely low.

Kiesling realized that synthetic biology offered a way to bypass the harvesting process altogether. By introducing the right genes into bacteria cells, he reasoned, the cells could turn into artemisinin-manufacturing machines, thus providing an abundant new source for the drug.

Getting there was tough. The team had to build an entirely new metabolic pathway into the cell, allowing it to process chemicals otherwise unknown to the cell.

Through trial-and-error, the team pasted together part of dozens of genes from several organisms into a custom DNA package. When they inserted the package into E. Coli, a bacteria commonly used in the lab to produce chemicals, it created a new pathway in the bacteria that allowed it to secrete artemisinin.

With more tinkering to increase efficacy, Kiesling and his team were able to bring up production by a factor of a million and reduce the drug’s price more than 10-fold.

Artemisinin was only the first step in a much larger program. The drug is a hydrocarbon, which belongs to a family of molecules often used to make biofuels. So why not use the same process to manufacture biofuels? By swapping out genes used to make artemisinin with those coding biofuel hydrocarbons, the team has already engineered multiple microbes capable of converting sugar to fuel.

Agriculture is another field poised to benefit from synthetic biology. Theoretically, we could take genes used to fix nitrogen from bacteria, put it into cells from our crops to completely alter their natural growth process. With the right combination of genes, we may be able to grow nutrition-packed crops — directed by an artificial genome — that require less water, land, energy and fertilizers.

Synthetic biology may even be used to produce completely new foods, such as flavorings created through fermentation with engineered yeast, or vegan cheeses and other animal-free milk products.

“We need to reduce carbon emissions and toxic inputs, use less land and water, combat pests, and increase soil fertility,” says Dr. Pamela Ronald, a professor at UC Davis. Synthetic biology may give us the tools to get there.

Re-creating Life

Practical applications aside, one of the ultimate goals of synthetic biology is to create a synthetic organism made exclusively from custom-designed DNA.

The main roadblock right now is technological. DNA synthesis is currently expensive, slow and prone to errors. Most existing techniques can only make DNA strands that are roughly 200 letters long, whereas genes are usually over ten times as long. The human genome contains roughly 20,000 genes that make proteins.

That said, costs for DNA synthesis have been rapidly dropping over the past decade.

According to Dr. Drew Endy, a geneticist at Stanford University, the cost of sequencing an individual letter has plummeted from $4 in 2003 to a mere 3 cents now. The estimated cost of printing all 3 billion letters of the human genome at the moment is a staggering $90 million, although that is expected to drop to $100,000 within 20 years if the trend continues.

An increasingly reasonable price tag has already opened doors to whole-genome synthesis.

Back in the 90s, Craig Venter, best known for his leading role in sequencing the human genome, began investigating the minimal set of genes required to make life. Together with colleagues at the Institute for Genomic Research, Venter removed genes from a bacterium Mycoplasma genitalium to identify those critical to life.

In 2008, Venter pieced together these “essential genes” and built the entire new “minimal” genome from a soup of chemicals using DNA synthesis.

Several years later, Venter transplanted the artificial genome into a second bacterium. The genes took over and “rebooted” the cell, allowing it to grow and self-replicate — the first living organism with a completely synthetic genome.

From Bacteria to Human

The new venture, if funded, would replicate Venter’s experiments using our own genome. Given that the human genome is nearly 5,000 times larger than Venter’s bacterium, it’s hard to say just how much more difficult the synthesis might be.

Even if that goal fails, however, the field is still bound to take a quantum leap forward. According to Dr. George Church, a leading geneticist at Harvard Medical School, the project could generate technological advances that improve our general ability to synthesize long strings of DNA — regardless of origin.

In fact, Church stressed that the project’s main goal is advancing technology.

But many are skeptical. According to Endy, who was invited to the meeting but decided to bow out, the project was originally named “HGP2: The Human Genome Synthesis Project,” and its primary goal was “to synthesize a complete human genome in a cell line within a period of 10 years.”

It’s perhaps not a surprise that news of the meeting caused a stir.

Regardless of its actual goals, the project raises the prospect of building custom-designed humans, or even semi-humans who have computers as parents.

The associated risks are easy to imagine and undoubtedly terrifying: how safe is it to directly manipulate or build life? How likely are accidents that unleash new organisms on an unprepared world? Who owns and has access to the technology? Would it breed new discrimination or further separate the 1% from the 99%?

“You can’t possibly begin to do something like this if you don’t have a value system in place that allows you to map concepts of ethics, beauty, and aesthetics onto our own existence,” says Endy.

“Given that human genome synthesis is a technology that can completely redefine the core of what now joins all of humanity together as a species, we argue that discussions of making such capacities real…should not take place without open and advance consideration of whether it is morally right to proceed,” he said.
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Breakthrough Holds Promise for Medicine: Synthetic Nanomotors Controlled Inside Live Human Cells

Breakthrough Holds Promise for Medicine: Synthetic Nanomotors Controlled Inside Live Human Cells | SynBioFromLeukipposInstitute | Scoop.it
Penn State University chemists and engineers have placed tiny synthetic motors inside live human cells, propelled them with ultrasonic waves, and steered them magnetically.
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Synthetic Human Genome Project

Synthetic Human Genome Project | SynBioFromLeukipposInstitute | Scoop.it
In a closed door meeting at Harvard on Tuesday nearly 150 scientists got together to organize the human synthetic genome project.
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Editorial: Engineering Synthetic Metabolons: From Metabolic Modeling to Rational Design of Biosynthetic Devices

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Engineering yeast shikimate pathway towards production of aromatics: rational design of a chassis cell using systems and synthetic biology - UQ eSpace

Aromatic building blocks are amongst the most important bulk feedstocks in the chemical industry. As these compounds are commonly derived from petrochemistry, obtaining them is becoming more and more a matter of costs and sustainability. Biochemistry gives rise to a wealth of compounds that can potentially replace current petroleum-based chemicals or be used for novel materials. The aromatic compounds para-aminobenzoic acid (pABA) and para-hydroxybenzoic acid (pHBA) and the aromatics derived compound cis,cis-muconic acid (ccMA) can be precursors for, but are not limited to, terephthalic or/and adipic acid. These are essential feedstocks for the production of PET and nylon. The three compounds can be derived from the shikimate pathway, an anabolic pathway leading to the biosynthesis of aromatic amino acids, present in certain prokaryotes and eukaryotes, including fungi. By combination of metabolic modelling with genetic engineering, a microbial production system based on the yeast Saccharomyces cerevisiae can be designed, which effectively channels flux into the target compounds.

In order to develop a competitive bio-based process, yields, titers and rates need to be maximized. While productivity or rates in a process can be altered using genetic engineering, carbon yield, and pathway feasibility are stoichiometrically and thermodynamically predetermined. Both limitations need to be considered when designing a microbial production system.

For formation of adipic acid and precursors many bio-based routes exists. More rational than just picking one for in vivo studies rather all available biochemical pathways were examined in silico using metabolic modelling. To compare theoretical yields and reaction thermodynamics an interface that allowed network-embedded thermodynamic analysis of elementary flux modes was developed. This allowed distinguishing between thermodynamically feasible and infeasible flux distributions. Feasible maximum theoretical product carbon yields were substantially different in E. coli and S. cerevisiae metabolic models and ranged from 32% to 92%. Further, many pathways appeared to be restricted by a thermodynamic equilibrium lying on the substrate side, some even infeasible.

The only routes that deliver significant product yields and were thermodynamically favoured were shikimate pathway based. Being currently of strong scientific interest, recent implementations of these pathways in E. coli and S. cerevisiae were evaluated and strain construction strategies optimized, using the concept of constrained minimal cut-sets. Especially in S. cerevisiae a single non-obvious knock-out target allowed coupling of growth to product formation; in particular, the deletion of the pyruvate kinase reaction resulted in a minimum yield constraint of 28%.

Though unique to shikimate pathway, this strategy is transferrable to other products which are derived from chorismate and also involve the formation of pyruvate as a by-product. This applies to pHBA. With further optimizations, the strategy was applied in vivo for the production of this compound in S. cerevisiae. As the pyruvate kinase knock-out entails a growth defect on glucose, a synthetic circuit was used, which allowed conditional knock-down and activation of the determined genetic modifications and by this dynamic control of the phenotype. Thus, production could be separated from growth and it could be proven that the in silico determined genetic intervention strategy holds valid in vivo, resulting in a 1.1 mM final product titer.

Further, production of pABA from shikimate pathway in S. cerevisiae was investigated and optimized: Several alleles from different yeast strains of the genes (ABZ1 and ABZ2) for pABA formation from chorismate were screened, using a strain genetically engineered to channel flux to chorismate. ABZ1 of AWRI1631 and ABZ2 of QA23 delivered the highest pABA production. To further increase production, the impact of carbon-source on product yield was investigated in silico using metabolic modelling. It was found that combined glycerol-ethanol was a superior carbon-source than glucose, glycerol or ethanol alone, especially when employed in molar ratios between 1:2 and 2:1. This could be confirmed in vivo with carbon yields reaching up to 2.64%. A fed-batch process on glycerol-ethanol delivered total aromatics titers as high as 1.73 mM.

It could be shown that feasibility and viability of adipic acid production greatly depends on the pathway and the organism chosen for engineering. Weaknesses of existing strain construction strategies for ccMA production could be identified and a radical optimization strategy was determined.

Inferences from metabolic modelling were proven experimentally for pHBA and pABA production. The obtained concentrations and yields are the highest in S. cerevisiae to date and among the highest in a microbial production system, underlining th
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Full speed ahead to the City on the Hill

Full speed ahead to the City on the Hill | SynBioFromLeukipposInstitute | Scoop.it
Landscape architects and ecological engineering scientists can design networks of self-sustaining and functioning natural ecosystems within the urban matrix. Urban parks, green roofs, and vegetation corridors will be more common and better connected to ensure the continuity of the natural ecosystem in the urban environment. The paradigm needs to shift from planting trees and building parks within the city to building the city around the natural environment.
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DIY Biology catches on with the UK’s first BioHackacthon

DIY Biology catches on with the UK’s first BioHackacthon | SynBioFromLeukipposInstitute | Scoop.it
The hackathon phenomena is knocking on biology’s door. The first UK Biohackathon will be held at the University of Cambridge in June, to learn and create with SynBio.

Hackathons are massively popular events in computer programming, joining people during a short period to create a hands-on project. The concept has now spread to biology – more specifically to synthetic biology.

This emerging area has a lot of potential, from biofuels to perfumes to healthcare. It already attracted a large bio-hacking community, the famous iGEM competitions and a specialized incubator.
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incaRNAfbinv: a web server for the fragment-based design of RNA sequences

In recent years, new methods for computational RNA design have been developed and applied to various problems in synthetic biology and nanotechnology. Lately, there is considerable interest in incorporating essential biological information when solving the inverse RNA folding problem. Correspondingly, RNAfbinv aims at including biologically meaningful constraints and is the only program to-date that performs a fragment-based design of RNA sequences. In doing so it allows the design of sequences that do not necessarily exactly fold into the target, as long as the overall coarse-grained tree graph shape is preserved. Augmented by the weighted sampling algorithm of incaRNAtion, our web server called incaRNAfbinv implements the method devised in RNAfbinv and offers an interactive environment for the inverse folding of RNA using a fragment-based design approach. It takes as input: a target RNA secondary structure; optional sequence and motif constraints; optional target minimum free energy, neutrality and GC content. In addition to the design of synthetic regulatory sequences, it can be used as a pre-processing step for the detection of novel natural occurring RNAs. The two complementary methodologies RNAfbinv and incaRNAtion are merged together and fully implemented in our web server incaRNAfbinv, available at http://www.cs.bgu.ac.il/incaRNAfbinv.
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Upgrading biomaterials with synthetic biological modules for advanced medical applications

One key aspect of synthetic biology is the development and characterization of modular biological building blocks that can be assembled to construct integrated cell-based circuits performing computational functions. Likewise, the idea of extracting biological modules from the cellular context has led to the development of in vitro operating systems. This principle has attracted substantial interest to extend the repertoire of functional materials by connecting them with modules derived from synthetic biology. In this respect, synthetic biological switches and sensors, as well as biological targeting or structure modules, have been employed to upgrade functions of polymers and solid inorganic material. The resulting systems hold great promise for a variety of applications in diagnosis, tissue engineering and drug delivery. This review reflects on the most recent developments and critically discusses challenges concerning in vivo functionality and tolerance that must be addressed to allow the future translation of such synthetic biology-upgraded materials from the bench to the bedside.
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What Innovative Investors Need to Know About Digital Biology

Is investing in synthetic biology worth the risk? 'Absolutely!' says Ryan Bethencourt of Berkeley Biolabs and Indie.Bio, an SOS Ventures backed biotech accelerator.
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Dopamine biosensor based on surface functionalized nanostructured nickel oxide platform | CodonOps

Dopamine biosensor based on surface functionalized nanostructured nickel oxide platform | CodonOps | SynBioFromLeukipposInstitute | Scoop.it
Dopamine biosensor based on surface functionalized nanostructured nickel oxide platform
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Enzyme assays using sensor arrays based on ion-selective carbon nanotube field-effect transistors

In the fields of clinical diagnostics and point-of-care diagnosis as well as food and environmental monitoring there is a high demand for reliable high-throughput, rapid and highly sensitive assays for a simultaneous detection of several analytes in complex and low-volume samples. Sensor platforms based on solution-processable electrolyte-gated carbon nanotube field-effect transistors (CNT-FETs) are a simple and cost-effective alternative for conventional assays. In this work we demonstrate a selective as well as direct detection of the products of an enzyme-substrate interaction, here the for metabolic processes important urea-urease system, with sensors based on spray-coated CNT-FETs. The selective and direct detection is achieved by immobilizing the enzyme urease via certain surface functionalization techniques on the sensor surface and further modifying the active interfaces with polymeric ion-selective membranes as well as pH-sensitive layers. Thereby, we can avoid the generally applied approach for a field-effect based detection of enzyme reactions via detecting changes in the pH value due to an on-going enzymatic reaction and directly detect selectively the products of the enzymatic conversion. Thus, we can realize a buffering-capacity independent monitoring of changes in the substrate concentration.
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