SynBioFromLeukipposInstitute

What's in a name?

Abstract
Defining an emerging field can be challenging. Nature Biotechnology asked 20 experts for their views on the term 'synthetic biology'.

Similar to other new and trendy fields, synthetic biology has been defined so loosely that it can seem like all things to all people. Traditional genetic or metabolic engineering has been rebranded as synthetic biology, often to take advantage of the hype cycle that fuels investor interest.

http://bit.ly/KRIhFE

Petya V Krasteva

"DNA's high chemical stability and programmable base-paired structure make it not only an ideal carrier of genetic information but also a promising material for engineering sophisticated nanodevices. Peng Yin and colleagues from the Wyss Institute at Harvard recently reported a robust bottom-up approach for the assembly of complex nanostructures in a single melting-annealing cycle from a mix of short synthetic DNAs.

The idea to use DNA as a scaffold in molecular engineering was conceived about 30 years ago when, inspired by M.C. Escher's woodwork 'Depth', crystallographer Ned Seeman reasoned that interlocking branched DNAs could be used as tiles to build carrier lattices for crystallization of target molecules. The concept launched the field of DNA nanotechnology and led to the first reports on the assembly of basic geometric shapes, tubes and lattices from DNA tiles. Nevertheless, complex nanoshapes remained largely out of reach, with experts sharing the intuition that imprecise ratios among the building tiles would lead to jammed assembly....."
http://bit.ly/LGoFnr

by

Alfred Pühler

*Synthetic antibody libraries*

by
Nelson B, Sidhu SS.
"Synthetic antibody libraries are constructed using designed synthetic DNA that facilitates the use of highly optimized human frameworks and enables the introduction of defined chemical diversity at positions that are most likely to contribute to antigen recognition. Using a relatively simple design based on a single human framework into which diversity is restricted to four complementarity-determining regions and two amino acids (tyrosine and serine), these synthetic antibody libraries are capable of generating specific antibodies against a diverse range of protein antigens. Moreover, by using the methods described here, more complex libraries can be constructed that are able to produce synthetic antibodies with affinities and specificities beyond the capacity of natural antibodies. Since these methods rely entirely upon standard supplies, equipment, and methods, construction of such libraries can be performed by any molecular biology laboratory."
http://1.usa.gov/KOPciF

by

Alkilany AM, Lohse SE, Murphy CJ.
"Since the late 1980s, researchers have prepared inorganic nanoparticles of many types-including elemental metals, metal oxides, metal sulfides, metal selenides, and metal tellurides-with excellent control over size and shape. Originally many researchers were primarily interested in exploring the quantum size effects predicted for such materials. Applications of inorganic nanomaterials initially centered on physics, optics, and engineering but have expanded to include biology. Many current nanomaterials can serve as biochemical sensors, contrast agents in cellular or tissue imaging, drug delivery vehicles, or even as therapeutics. In this Account we emphasize that the understanding of how nanomaterials will function in a biological system relies on the knowledge of the interface between biological systems and nanomaterials, the nano-bio interface. Gold nanoparticles can serve as excellent standards to understand more general features of the nano-bio interface because of its many advantages over other inorganic materials. The bulk material is chemically inert, and well-established synthetic methods allow researchers to control its size, shape, and surface chemistry. Gold's background concentration in biological systems is low, which makes it relatively easy to measure it at the part-per-billion level or lower in water. In addition, the large electron density of gold enables relatively simple electron microscopic experiments to localize it within thin sections of cells or tissue. Finally, gold's brilliant optical properties at the nanoscale are tunable with size, shape, and aggregation state and enable many of the promising chemical sensing, imaging, and therapeutic applications. Basic experiments with gold nanoparticles and cells include measuring the toxicity of the particles to cells in in vitro experiments. The species other than gold in the nanoparticle solution can be responsible for the apparent toxicity at a particular dose. Once the identity of the toxic agent in nanoparticle solutions is known, researchers can employ strategies to mitigate toxicity. For example, the surfactant used at high concentration in the synthesis (0.1 M) of gold nanorods remains on their surface in the form of a bilayer and can be toxic to certain cells at 200 nM concentrations. Several strategies can alleviate the toxic response. Polyelectrolyte layer-by-layer wrapping can cover up the surfactant bilayer, or researchers can exchange the surfactant with chemically similar molecules. Researchers can also replace the surfactant with a biocompatible thiol or use a polymerizable surfactant that can be "stitched" onto the nanorods and reduce its lability. In all these cases, however, proteins or other molecules from the cellular media cover the engineered surface of the nanoparticles, which can drastically change the charges and functional groups on the nanoparticle surface."

http://1.usa.gov/QjR2Yv

by

Carrasquilla C, Lau PS, Li Y, Brennan JD.
"Structure-switching, fluorescence-signaling DNA and RNA aptamers have been reported as highly versatile molecular recognition elements for biosensor development. While structure-switching DNA aptamers have been utilized for solid-phase sensing, equivalent RNA aptamers have yet to be successfully utilized in solid-phase sensors due to their lack of chemical stability and susceptibility to nuclease attack. In this study, we examined entrapment into sol-gel derived organic-inorganic composite materials as a platform for immobilization of structure-switching fluorescence-signaling RNA aptamer reporters, using both the synthetic theophylline- and naturally occurring thiamine pyrophosphate-binding RNA aptamers as test cases. Structure-switching versions of both aptamers were entrapped into a series of sol-gel derived composites, ranging from highly polar silica to hydrophobic methylsilsesquioxane-based materials, and the target-binding and signaling capabilities of these immobilized aptamers were assessed relative to solution. Both immobilized aptamers demonstrated sensitivity and selectivity similar to that of free aptamers when entrapped in a composite material derived from 40% (v/v) methyltrimethoxysilane/tetramethoxysilane. Importantly, this material also conferred protection from nuclease degradation and imparted long-term chemical stability to the RNA reporter systems. Given the versatility of sol-gel entrapment for development of biosensors, microarrays, bioaffinity columns, and other devices, this entrapment method should provide a useful platform for numerous solid-phase RNA aptamer-based devices."
http://1.usa.gov/MWGz0F

by

Christina Agapakis

How would you customize your garden?

http://bit.ly/MOsRkc

by

Wieland M, Fussenegger M.
"In most approaches, personalized medicine requires time- and cost-intensive characterization of an individual's genetic background in order to achieve the best-adapted therapy. For this purpose, cell-based drug delivery offers a promising alternative. In particular, synthetic biology has introduced the vision of cells being programmable therapeutic production facilities that can be introduced into patients. This review highlights the progress made in synthetic biology-based cell engineering toward advanced drug delivery entities. Starting from basic one-input responsive transcriptional or post-transcriptional gene control systems, the field has reached a level on which cells can be engineered to detect cancer cells, to obtain control over T-cell proliferation, and to restore blood glucose homeostasis upon blue light illumination. Furthermore, a cellular implant was developed that detects blood urate level disorders and acts accordingly to restore homeostasis while another cellular implant was engineered as an artificial insemination device that releases bull sperm into bovine ovarian only during ovulation time by recording endogenous luteinizing hormone levels. Soon, the field will reach a stage at which cells can be reprogrammed to detect multiple metabolic parameters and self-sufficiently treat any disorder connected to them."
http://1.usa.gov/NkYEH3

"In treating cancer patients, one problem researchers face is how to deliver as much of the therapeutics as possible directly to cancer cells, while sparing normal cells. Many are working on various delivery systems using nanotech, synthetic biology approaches, RNA interference, and more. Now, a group at Johns Hopkins University led by Samuel Denmeade is taking inspiration from nature, says New Scientist's Hannah Krakauer. In its study published in Science Translational Medicine, the group describes how it engineered an analogue of thapsigargin, a toxic substance found in the flowering Thapsia garganica plant — nicknamed the "death carrot" by the ancient Greeks because of its toxicity to sheep and cattle. "Thapsigargin typically works by passing through cell membranes and shutting down calcium pumps — essential for cell survival — on the inside of cells," Krakauer says. By adding an extra peptide chain to the toxin, Denmeade's team changed it enough to keep it from entering cells until is encounters an enzyme called PSMA, which is found the surface of prostate cancer cells. "PSMA cleaves the extra chain off the toxin, setting it free to do its devastating business," Krakauer adds.

Further, the toxin doesn't just kill cancer cells that are undergoing growth, but can also kill dormant tumor cells, and non-cancerous cells used by tumors to help them grow. "You can envision it as a grenade," Denmeade tells Krakauer. "One guy pulls the pin, but it kills all the guys standing around." As the researchers' animal trials were successful, they are now moving to phase I clinical trials in prostate cancer, Krakauer adds."
http://bit.ly/Mcpfuv

Ref
Engineering a Prostate-Specific Membrane Antigen–Activated Tumor Endothelial Cell Prodrug for Cancer Therapy
by
Samuel R. Denmeade1,*, Annastasiah M. Mhaka1, D. Marc Rosen1, W. Nathaniel Brennen1, Susan Dalrymple1, Ingrid Dach2, Claus Olesen2, Bora Gurel1, Angelo M. DeMarzo1, George Wilding3, Michael A. Carducci1, Craig A. Dionne4, Jesper V. Møller2,5, Poul Nissen2,6, S. Brøgger Christensen7 and John T. Isaacs1
"Heterogeneous expression of drug target proteins within tumor sites is a major mechanism of resistance to anticancer therapies. We describe a strategy to selectively inhibit, within tumor sites, the function of a critical intracellular protein, the sarcoplasmic/endoplasmic reticulum calcium adenosine triphosphatase (SERCA) pump, whose proper function is required by all cell types for viability. To achieve targeted inhibition, we took advantage of the unique expression of the carboxypeptidase prostate-specific membrane antigen (PSMA) by tumor endothelial cells within the microenvironment of solid tumors. We generated a prodrug, G202, consisting of a PSMA-specific peptide coupled to an analog of the potent SERCA pump inhibitor thapsigargin. G202 produced substantial tumor regression against a panel of human cancer xenografts in vivo at doses that were minimally toxic to the host. On the basis of these data, a phase 1 dose-escalation clinical trial has been initiated with G202 in patients with advanced cancer."
http://bit.ly/NWMKXm

We are LIVE! Win cash and global bragging rights for developing Open Source tools for Synthetic and DIY Biology at

http://sprize.synbiota.com/

Genome Editing with CompoZr Custom Zinc Finger Nucleases (ZFNs)

by
Hansen K, Coussens MJ, Sago J, Subramanian S, Gjoka M, Briner D.
"Genome editing is a powerful technique that can be used to elucidate gene function and the genetic basis of disease. Traditional gene editing methods such as chemical-based mutagenesis or random integration of DNA sequences confer indiscriminate genetic changes in an overall inefficient manner and require incorporation of undesirable synthetic sequences or use of aberrant culture conditions, potentially confusing biological study. By contrast, transient ZFN expression in a cell can facilitate precise, heritable gene editing in a highly efficient manner without the need for administration of chemicals or integration of synthetic transgenes. Zinc finger nucleases (ZFNs) are enzymes which bind and cut distinct sequences of double-stranded DNA (dsDNA). A functional CompoZr ZFN unit consists of two individual monomeric proteins that bind a DNA "half-site" of approximately 15-18 nucleotides (see Figure 1). When two ZFN monomers "home" to their adjacent target sites the DNA-cleavage domains dimerize and create a double-strand break (DSB) in the DNA.(1) Introduction of ZFN-mediated DSBs in the genome lays a foundation for highly efficient genome editing. Imperfect repair of DSBs in a cell via the non-homologous end-joining (NHEJ) DNA repair pathway can result in small insertions and deletions (indels). Creation of indels within the gene coding sequence of a cell can result in frameshift and subsequent functional knockout of a gene locus at high efficiency.(2) While this protocol describes the use of ZFNs to create a gene knockout, integration of transgenes may also be conducted via homology-directed repair at the ZFN cut site. The CompoZr Custom ZFN Service represents a systematic, comprehensive, and well-characterized approach to targeted gene editing for the scientific community with ZFN technology. Sigma scientists work closely with investigators to 1) perform due diligence analysis including analysis of relevant gene structure, biology, and model system pursuant to the project goals, 2) apply this knowledge to develop a sound targeting strategy, 3) then design, build, and functionally validate ZFNs for activity in a relevant cell line. The investigator receives positive control genomic DNA and primers, and ready-to-use ZFN reagents supplied in both plasmid DNA and in-vitro transcribed mRNA format. These reagents may then be delivered for transient expression in the investigator's cell line or cell type of choice. Samples are then tested for gene editing at the locus of interest by standard molecular biology techniques including PCR amplification, enzymatic digest, and electrophoresis. After positive signal for gene editing is detected in the initial population, cells are single-cell cloned and genotyped for identification of mutant clones/alleles."
http://1.usa.gov/MU6Kap

by

O'Brien EL, Itallie EV, Bennett MR.
"Genetic oscillators have long held the fascination of experimental and theoretical synthetic biologists alike. From an experimental standpoint, the creation of synthetic gene oscillators represents a yardstick by which our ability to engineer synthetic gene circuits can be measured. For theorists, synthetic gene oscillators are a playground in which to test mathematical models for the dynamics of gene regulation. Historically, mathematical models of synthetic gene circuits have varied greatly. Often, the differences are determined by the level of biological detail included within each model, or which approximation scheme is used. In this review, we examine, in detail, how mathematical models of synthetic gene oscillators are derived and the biological processes that affect the dynamics of gene regulation."
http://1.usa.gov/MP5ntI

"Scientists may have hit gold in their fight against dengue. They have located a human antibody that can neutralise and kill its virus within two hours.

Significantly, they have also identified a way to reproduce this antibody in large quantities, potentially opening the door to a cure for dengue infected patients.

The symptoms of dengue are sudden-onset fever, headache (located behind the eyes), muscle and joint pains, and a rash. The alternative name for dengue, ‘break-bone fever,’ which comes from the associated muscle and joint pains.

This discovery was made by a combined team from the National University of Singapore (NUS) Yong Loo Lin School of Medicine, Duke-NUS Graduate Medical School and the Defence Medical & Environmental Research Institute, all based in Singapore."
http://bit.ly/MQ1NB7

"Jason Bobe serves as the Executive Director of PersonalGenomes.org and Director of Community for the Personal Genome Project based out of George Church’s lab at Harvard Medical School. The Personal Genome Project seeks to encourage the thoughtful development of personal genomics technology and practices by building frameworks for prototyping and evaluation at increasing scales.

Jason is co-founder of DIYbio.org, an organization that aims to help make biology a worthwhile pursuit for citizen scientists and amateur biologists. DIYbio is fast becoming the organizational hub for amateur biologists worldwide, uniting the movement’s participants through its website, online forums, blog and local chapters.
He has also worked as a Business Development Consultant for OpenWetWare.org, as Director of Business Development at DNA Direct, and as an independent consultant. He has attended both the University of Colorado at Boulder and the Kelley School of Business at Indiana University. He is interested in how emerging biotechnologies and the web are redefining the relationships between scientific research communities, communities from the general public, and the network of actors in-between..."

http://bit.ly/KVtXNx

an interesting list of unpublished papers

and some free downloads
http://bit.ly/nd7L2K