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Scooped by Dr. Stefan Gruenwald!

Why De-Extinction of Birds is a Challenge – The Passenger Pidgeon Case

Why De-Extinction of Birds is a Challenge – The Passenger Pidgeon Case | Amazing Science |

Birds are a huge challenge for de-extinction for two big reasons. The first is because less genomic research has been performed on birds than on mammals (but reptiles, amphibians, fish, invertebrates and plants are even less understood). We don’t know how precisely how the majority of gene pathways in birds work on the cellular levels and up.

Also, birds have no uterus. The reason that the absence of a uterus is a problem for cloning relates to how cloning is done. When you take the nucleus out of an egg cell you kill that cell, it is completely dead. Even after you put a new nucleus in it, the cell is still dead. You have to bring the cell back to life, just like when you shock someone’s heart into beating again. You run electricity through the newly cloned cell to get it to divide. The problem here is that you have to keep stimulating cell division for many generations, up to several hundred and even a few thousand cells before the embryo will develop on its own without assistance. Therefore you cannot take a single cloned cell and implant it into an ovary, oviduct, uterus or any reproductive organ and get it to grow – you have to grow it in the lab and then implant a partially developed embryo. This is okay in a uterus because the embryo implants and develops in a fixed place. In a bird, the embryo is in constant motion within the female’s body – literally tumbling down the oviduct as the oviduct coats the eggshell around the embryo. To implant a cloned embryo one would have to take out the developing embryo from within a developing hard shelled egg within the female’s body and replace it with the cloned embryo – and hope that the embryo integrates into the yolk of the egg and that all the puncturing doesn’t deform the egg or harm the female. So you can see it’s very very tricky.

Are there ways to introduce an extinct bird’s genetics into an embryo without cloning? You can introduce cells into the embryo, which will integrate and create a chimeric bird – a bird that has a patchwork of tissues made of cells of both the original embryo and the cells that were introduced. This can be done after the egg is laid, avoiding tampering with the mother’s internal organ systems. The problem for de-extinction is that adult stem cells (or induced Pluripotent Stem cells, iPCs) cannot contribute to the germ line, only Embryonic stem cells can contribute to the germ line. We can’t easily use embryonic stem cells to recreate the passenger pigeon genome. After as few as seven days in a lab culture, embryonic stem cells have undergone enough cell division to be adult stem cells, and lose the ability to become germ cells. A process to use embryonic stem cells would require introducing a mutation to a band-tailed pigeon embryonic stem cell in less than a matter of a few days, then put it into an embryo and hatch a chimera. This would then require hundreds to even thousands of generations of chimeric birds until we have a passenger pigeon. It would be far more efficient to introduce the thousands of mutations in cell lines, then create a bird. But by the time all the mutations were added, the cells would be adult stem cells. You could make as many chimeras as you want from these “de-extinct” stem cells, but they would never form a breeding line. This does not mean that stem cells cannot become germ cells under experimental conditions, what this means is that they do not naturally become germ cells when placed inside a developing bird embryo. It may be possible in the future to program iPSCs to become germ cells, but currently this is not possible.

Further reading: The Mammoth Cometh (NY Times)

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New live-cell printing technology improves on inkjet printing

New live-cell printing technology improves on inkjet printing | Amazing Science |

A new way to print living cells onto any surface and in almost any shape has been developed by researchers led by Houston Methodist Research Institute nanomedicine faculty member Lidong Qin. Unlike a similar inkjet printing process, almost all cells survive.

The new process, called Block-Cell-Printing (BloC-Printing), produces 2-D cell arrays in half an hour, prints the cells as close together as 5 microns (most animal cells are 10 to 30 microns wide), and allows the use of many different cell types.

“Cell printing is used in so many different ways now — for drug development and in studies of tissue regeneration, cell function, and cell-cell communication,” Qin said. “Such things can only be done when cells are alive and active. A survival rate of 50 to 80 percent is typical as cells exit the inkjet nozzles. “By comparison, we are seeing close to 100 percent of cells in BloC-Printing survive the printing process.”

BloC-Printing manipulates microfluidic physics to guide living cells into hook-like traps in the silicone mold. Cells flow down a column in the mold, past trapped cells to the next available slot, eventually creating a line of cells in a grid.

The position and spacing of the traps and the shape of the channel navigated by the cells is fully configurable during the mold’s creation. When the mold is lifted away, the living cells remain behind, adhering to the growth medium or other substrate, in prescribed formation.

The researchers also printed a grid of brain cells and gave the cells time to form synaptic and autaptic junctions. “The cell junctions we created may be useful for future neuron signal transduction and axon regeneration studies,” Qin said. “Such work could be helpful in understanding Alzheimer’s disease and other neurodegenerative diseases.”

While it is too early to predict the market cost of BloC-Printing, Qin said the materials of a single BloC mold cost about $1 (US). After the mold has been fabricated and delivered, a researcher only needs a syringe, a carefully prepared suspension of living cells, a Petri dish, and a steady hand, Qin said. Inkjet cell printers can cost between $10,000 and $200,000.

“BloC-Printing can be combined with molecular printing for many types of drug screening, RNA interference, and molecule-cell interaction studies,” he said. “We believe the technology has big potential.” While the fidelity of BloC-Printing is high, Qin said inkjet printing remains faster, and BloC-Printing cannot yet print multi-layer structures as inkjetting can.

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Researchers use Google's exacycle cloud computing platform to simulate key drug receptor

Researchers use Google's exacycle cloud computing platform to simulate key drug receptor | Amazing Science |

Roughly 40 percent of all medications act on cells' G protein-coupled receptors. One of these receptors, beta 2 adrenergic receptor site (B2AR), naturally transforms between two base configurations; knowing the precise location of each of approximately 4,000 atoms is crucial for ensuring a snug fit between it and the drug.

Now, researchers at Stanford and Google have conducted an unprecedented, atom-scale simulation of the receptor site's transformation, a feat that could have significant impact on drug design. This is the first scientific project to be completed using Google Exacycle's cloud computing platform, which allows scientists to crunch big data on Google's servers during periods of low network demand.

The study was published in the January issue of Nature Chemistry.

As a type of GPCR, the B2AR is a molecule that sits within the membrane of most cells. Various molecules in the body interact with the receptor's exterior, like two hands shaking, to trigger an action inside the cell.

"GPCRs are the gateway between the outside of the cell and the inside," said co-author Vijay Pande, PhD, professor of chemistry and a senior author of the study. "They're so important for biology, and they're a natural, existing signaling pathway for drugs to tap into."

Lead authors of the study were former postdoctoral scholar Kai Kohlhoff, PhD, and current postdoctoral scholars Diwakar Shukla, PhD, and Morgan Lawrenz, PhD.

Roughly half of all known drugs—including pharmaceuticals and naturally occuring molecules, such as caffeine —target some GPCR, and many new medications are being designed with these receptor sites in mind. Brian Kobilka, professor of molecular and cellular physiology at Stanford, was awarded the 2012 Nobel Prize in Chemistry for his role in discovering and understanding GPCRs.

Traditionally, maps that detail each atom of GPCRs and other receptors are created through a technique called X-ray crystallography. The technique is industry standard, but it can only visualize a molecule in its resting state; receptors naturally change configurations, and their intermediate forms might also have medical potential.

When developing a drug, scientists will often run a computer program, known as a docking program, that predicts how well the atomic structure of a proposed drug will fit into the known receptor.

In the case of GPCRs, for example, the X-ray crystallography techniques have detailed their "on" and "off" configurations; many medications have been specifically designed to fit into these sites. Scientists expect, however, that other fruitful configurations exist. Many drugs engage with GPCR sites, even though computational models suggest that they don't fit either of the two defined reaction site configurations.

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Scientists move closer to stem cell cure for type 1 diabetes

Scientists move closer to stem cell cure for type 1 diabetes | Amazing Science |

Researchers say they have reversed equivalent of type 1 diabetes in mice using stem cell transplants.

Researchers in California report that they have reversed the equivalent of type 1 diabetes in mice through transplants of stem cells. Their experiments have replaced cells in the pancreas damaged by the disease that are unable to make insulin. Without insulin, the body has difficulty absorbing sugars such as glucose from the blood. The disease usually first shows in childhood or early adulthood and used to be a killer, but glucose levels can now be monitored and regulated with insulin injections.

Scientists have long wanted to try to replace the damaged ß-cells that normally produce insulin. This has been one of the prime targets of stem cell experiments. But until now, it has proved difficult, partly because mature ß-cells do not readily regenerate.

Writing in the journal Cell Stem Cell, scientists at the Gladstone Institutes in San Francisco describe how they took a step back and collected skin cells, called fibroblasts, from laboratory mice. Then, by treating the fibroblasts with a unique "cocktail" of molecules and reprogramming factors, they transformed the cells into endoderm-like cells. Endoderm cells are a type of cell found in the early embryo, and which eventually mature into the body's major organs – including the pancreas.

"Using another chemical cocktail, we then transformed these endoderm-like cells into cells that mimicked early pancreas-like cells, which we called PPLCs," said the Gladstone postdoctoral scholar Ke Li, the paper's lead author. "Our initial goal was to see whether we could coax these PPLCs to mature into cells that, like ß-cells, respond to the correct chemical signals and – most importantly – secrete insulin. And our initial experiments, performed in a petri dish, revealed that they did."

The team then injected these cells into mice that had been genetically modified to have high glucose levels, mimicking the type 1 diabetes condition in humans. "Importantly, just one week post-transplant, the animals' glucose levels started to decrease, gradually approaching normal levels," said Li. "And when we removed the transplanted cells, we saw an immediate glucose spike, revealing a direct link between the transplantation of the PPLCs and reduced hyperglycemia."

Eight weeks after the transplantation, the scientists found that the pancreas-like cells had turned into the real thing – fully functional insulin-secreting ß-cells had developed in the mice.

The team says this is proof of principle, which one day might be used to cure type 1 diabetes in humans. "I am particularly excited about the prospect of translating these findings to the human system," said Matthias Hebrok, one of the study's authors and director of the UCSF Diabetes Center. "Most immediately, this technology in human cells could significantly advance our understanding of how inherent defects in ß-cells result in diabetes, bringing us notably closer to a much-needed cure."

Michelle Milici's curator insight, November 4, 2014 3:14 PM

This is a recent possibility that could change so many lives. A potential cure for type 1 diabetes could also lead to a cure for type 2 diabetes

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Crystallographic elucidation of CRISPR-Cas9 endonuclease complex reveals RNA-mediated conformational changes

Crystallographic elucidation of CRISPR-Cas9 endonuclease complex reveals RNA-mediated conformational changes | Amazing Science |

The potential is there for bacteria and other microbes to be genetically engineered to perform a cornucopia of valuable goods and services, from the production of safer, more effective medicines and clean, green, sustainable fuels, to the clean-up and restoration of our air, water and land. Cells from eukaryotic organisms can also be modified for research or to fight disease. To achieve these and other worthy goals, the ability to precisely edit the instructions contained within a target's genome is a must. A powerful new tool for genome editing and gene regulation has emerged in the form of a family of enzymes known as Cas9, which plays a critical role in the bacterial immune system. Cas9 should become an even more valuable tool with the creation of the first detailed picture of its three-dimensional shape by researchers with the Lawrence Berkeley National Laboratory (Berkeley Lab) and the University of California (UC) Berkeley.

Biochemist Jennifer Doudna and biophysicist Eva Nogales, both of whom hold appointments with Berkeley Lab, UC Berkeley, and the Howard Hughes Medical Institute (HHMI), led an international collaboration that used x-ray crystallography to produce 2.6 and 2.2 angstrom (Å) resolution crystal structure images of two major types of Cas9 enzymes. The collaboration then used single-particle electron microscopy to reveal how Cas9 partners with its guide RNA to interact with target DNA. The results point the way to the rational design of new and improved versions of Cas9 enzymes for basic research and genetic engineering.

Cas9 is a family of RNA-guided bacterial endonucleases employed by Type II CRISPR systems to recognize and cleave double-stranded DNA at site-specific sequences. Genetic engineers have begun harnessing Cas9 for genome editing and gene regulation in many eukaryotic organisms. However, despite the successes to date, the technology has yet to reach its full potential because until now the structural basis for guide RNA recognition and DNA targeting by Cas9 has been unknown.

What has been a major puzzle in the CRISPR–Cas field is how Cas9 and similar RNA-guided complexes locate and recognize matching DNA targets in the context of an entire genome, the classic needle in a haystack problem," says Samuel Sternberg, lead author of the Nature paper and a member of Doudna's research group. "All of the scientists who are developing RNA-programmable Cas9 for genome engineering are relying on its ability to target unique 20-base-pair long sequences inside the cell. However, if Cas9 were to just blindly bind DNA at random sites across a genome until colliding with its target, the process would be incredibly time-consuming and probably too inefficient to be effective for bacterial immunity, or as a tool for genome engineers. Our study shows that Cas9 confines its search by first looking for PAM sequences. This accelerates the rate at which the target can be located, and minimizes the time spent interrogating non-target DNA sites."

Now, several scientists addressed this lack of detailed knowledge about Cas9 by first solving the three-dimensional crystal structures of two Cas9 proteins, representing large and small versions, from Streptococcus pyogenes (SpyCas) and Actinomyces naeslundii (AnaCas9) respectively. Using protein crystallography beamlines at Berkeley Lab's Advanced Light Source and the Paul Scherer Institute's Swiss Light Source, the collaboration discovered that despite significant differences outside of their catalytic domains, all members of the Cas9 family share the same structural core. The high resolution images showed this core to feature a clam-shaped architecture with two major lobes - a nuclease domain lobe and an alpha-helical lobe. Both lobes contained conserved clefts that become functional in nucleic acid binding.

"Our understanding of Cas9's structure was not complete with only the x-ray data because the protein in the crystals had been trapped in a state without its associated guide RNA," says Sam Sternberg, a member of Doudna's research group and a co-author of the Science paper. "Understanding how RNA-guided Cas9 targets matching DNA sequences for genome engineering and how this reaction and its specificity might be improved required an understanding of how the shape of Cas9 changes when it interacts with guide RNA, and when a matching DNA target sequence is bound."

The collaboration employed negative-staining electron microscopy to visualize the Cas9 protein bound to either guide RNA, or both RNA and target DNA. The structures revealed that the guide RNA binding structurally activates Cas9 by creating a channel between the two main lobes of the protein that functions as the DNA-binding interface.

"Our single particle electron microscopy analysis reveals the importance of guide-RNA for the conversion of Cas9 into a structurally-activated state," says David Taylor, a joint member of Doudna's and Nogales's research groups and another co-author of the Science paper. "The results underline that, in addition to sequence complementarity, other features of the guide-RNA must be considered when employing this technology."

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Capturing ten-color ultrasharp images of synthetic DNA structures resembling numerals 0 to 9

Capturing ten-color ultrasharp images of synthetic DNA structures resembling numerals 0 to 9 | Amazing Science |

A new microscopy method could enable scientists to generate snapshots of dozens of different biomolecules at once in a single human cell, a team from the Wyss Institute of Biologically Inspired Engineering at Harvard University reported Sunday in Nature Methods.

Such images could shed light on complex cellular pathways and potentially lead to new ways to diagnose disease, track its prognosis, or monitor the effectiveness of therapies at a cellular level.

Cells often employ dozens or even hundreds of different proteins and RNA molecules to get a complex job done. As a result, cellular job sites can resemble a busy construction site, with many different types of these tiny cellular workers coming and going. Today's methods typically only spot at most three or four types of these tiny workers simultaneously. But to truly understand complex cellular functions, it's important to be able to visualize most or all of those workers at once, said Peng Yin, Ph.D., a Core Faculty member at the Wyss Institute and Assistant Professor of Systems Biology at Harvard Medical School.

To capture ultrasharp images of biomolecules, they had to overcome laws of physics that stymied microscopists for most of the last century. When two objects are closer than about 200 nanometers apart — about one five-hundredth the width of a human hair — they cannot be distinguished using a traditional light microscope: the viewer sees one blurry blob where in reality there are two objects.

Since the mid-1990s, scientists have developed several ways to overcome this problem using combinations of specialized optics, special fluorescent proteins or dyes that tag cellular components.

Ralf Jungmann, Ph.D., now a Postdoctoral Fellow working with Yin at the Wyss Institute and Harvard Medical School, helped develop one of those super-resolution methods, called DNA-PAINT, as a graduate student. DNA-PAINT can create ultrasharp snapshots of up to three cellular workers at once by labeling them with different colored dyes.

To visualize cellular job sites with crews of dozens of cellular workers, Yin's team, including Jungmann, Maier Avendano, M.S., a graduate student at Harvard Medical School, and Johannes Woehrstein, a postgraduate research fellow at the Wyss Institute, modified DNA-PAINT to create a new method called Exchange-PAINT.

Exchange-PAINT relies on the fact that DNA strands with the correct sequence of letters, or nucleotides, bind specifically to partner strands with complementary sequences. The researchers label a biomolecule they want to visualize with a short DNA tag, then add to the solution a partner strand carrying a fluorescent dye that lights up only when the two strands pair up. When that partner strand binds the tagged biomolecule, it lights up, then lets go, causing the biomolecule to "blink" at a precise rate the researchers can control. The researchers use this blinking to obtain ultrasharp images.

To test Exchange-PAINT, the researchers created 10 unique pieces of folded DNA, or DNA origami, that resembled the numerals 0 through 9. These numerals could be resolved with less than 10 nanometers resolution, or one-twentieth of the diffraction limit.

The team was able to use Exchange-PAINT to capture clear images of the 10 different types of miniscule DNA origami structures in one image. They also used the method to capture detailed, ultrasharp images of fixed human cells, with each color tagging an important cellular component — microtubules, mitochondria, Golgi apparatus, or peroxisomes.

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Genetically modified purple tomatoes heading for shops

Genetically modified purple tomatoes heading for shops | Amazing Science |

The prospect of genetically modified (GM) purple tomatoes reaching the shelves has come a step closer. Their dark pigment is intended to give tomatoes the same potential health benefits as fruit such as blueberries.

Developed in Britain, large-scale production is now under way in Canada with the first 1,200 liters of purple tomato juice ready for shipping. The pigment, known as anthocyanin, is an antioxidant which studies on animals show could help fight cancer.

Scientists say the new tomatoes could improve the nutritional value of everything from ketchup to pizza topping. The tomatoes were developed at the John Innes Centre in Norwich where Prof Cathie Martin hopes the first delivery of large quantities of juice will allow researchers to investigate its potential.

"With these purple tomatoes you can get the same compounds that are present in blueberries and cranberries that give them their health benefits - but you can apply them to foods that people actually eat in significant amounts and are reasonably affordable," she said.

The tomatoes are part of a new generation of GM plants designed to appeal to consumers - the first types were aimed specifically at farmers as new tools in agriculture. The purple pigment is the result of the transfer of a gene from a snapdragon plant - the modification triggers a process within the tomato plant allowing the anthocyanin to develop.

Although the invention is British, Prof Martin says European Union restrictions on GM encouraged her to look abroad to develop the technology. Canadian regulations are seen as more supportive of GM and that led to a deal with an Ontario company, New Energy Farms, which is now producing enough purple tomatoes in a 465 square meter (5,000sq ft) greenhouse to make 2,000 liters (440 gallons) of juice. The first 1,200 liters are due to be shipped to Norwich shortly - and because all the seeds will have been removed, there is no genetic material to risk any contamination.

Laura E. Mirian, PhD's curator insight, February 1, 2014 2:31 PM


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Researchers Develop DNA-based Method For Authenticating Premium Chocolate

Researchers Develop DNA-based Method For Authenticating Premium Chocolate | Amazing Science |
The taste of a rich, thick morsel of luxurious premium chocolate can be the ultimate experience for some people. But how do you know you are getting what you paid for? Until now, chocolate connoisseurs relied on just their taste buds.

A new study, published in the American Chemical Society’s (ACM) Journal of Agricultural and Food Chemistry, reports that a method to authenticate the varietal purity and origin of cacao beans — the source of chocolate’s main ingredient, cocoa—has been developed for the first time.

Lower-quality cacao beans often get mixed in with premium varieties on their way to becoming chocolate bars, truffles, sauces and liqueurs, said Dapeng Zhang, postdoctoral researcher at the National Center for Biotechnology Information. However, the stakes for policing the chocolate industry are high because it’s a multi-billion dollar global enterprise. In some areas, being a chocolatier is as much an art form as a business. Conservation also plays a role in knowing whether products are truly what the confectioners claim them to be in that the ability to authenticate rare varieties would encourage growers to maintain cacao biodiversity rather than depend on the most abundant and easiest to grow plants.

Using genetic testing, researchers have discovered ways to verify the authenticity of many other crops, such as cereals, fruits, olives, tea and coffee. However, these methods are not suitable for cacao beans, leading Zhang and his team to address the challenge of finding alternative methods.

The team applied the most recent developments in cacao genomics to identify a small set of DNA markers known as SNPs (pronounced “snips”). These SNPs make up unique fingerprints of different cacao species. The team found that the technique works on single cacao beans and can also be scaled up to handle large samples quickly.

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Storing data as DNA – as easy as ACTG?

Storing data as DNA – as easy as ACTG? | Amazing Science |

iSGTW recently interviewed Ewan Birney, associate director of the European Bioinformatics Institute (EBI), regarding his keynote talk at the EUDAT 2nd Conference. In this interview, Birney raised the exciting prospect of using DNA as an organic data storage device. But could DNA storage really replace tapes and hard disks for long-term preservation of data?

Deinococcus radiodurans, for example, a bacterial species adapted to survive in extreme conditions, was chosen by Wong and his colleagues to be the host due to its ability to quickly repair spontaneously arising mutations. Unlike hard drives or magnetic tapes, which are vulnerable to physical damage, data stored in bacteria could survive numerous natural disasters and be safely passed on to future generations.

However, the heterologous (artificially inserted) DNA could make the bacterial genome unstable, believesGeoff Baldwin, reader in biochemistry at Imperial College London, UK. “Bacteria are highly evolved organisms with relatively minimal genomes”, says Baldwin. “There is always the issue that maintaining large quantities of heterologous DNA will exert a fitness burden that will favor loss of the additional DNA, which does not bode well for the use of bacteria as a mass data storage device.”

While mutation rates are relatively low (approximately 1 base every 10,000 generations), bacteria's fast replication rate could make long-term data storage problematic. Another issue is that if the inserted DNA is similar to that of the host bacteria, it could interfere with its normal cellular processes. “Ultimately this means that there is not complete freedom to insert any sequence,” says Baldwin. “This can be overcome to some extent depending on the method chosen to encode information, but there always remains the possibility for instability due to unexpected consequences.”

While data storage in bacteria isn't yet sufficiently developed to be used for mass storage, using ‘naked’ DNA could be a more promising alternative. Mammoths and Neanderthals have been found preserved for thousands of years with DNA sequences intact, showing that living cells are not required for DNA itself to remain an efficient, stable data-storage means. Naked DNA is easier to use than bacteria as it doesn't require genetic manipulations to safely insert it into a host.

Birney was part of a team that encoded a record-breaking 700 kilobytes of unique data — including all 154 of Shakespeare's sonnets — into naked DNA and retrieved it with 99% accuracy. Translating binary data into a 4-base system often results in long sequences of identical bases, which have a tendency to be misread by DNA-sequencing machines, ultimately degrading the information of the original message. The team came up with an ingenious system that allowed them to encode data with such high fidelity. They used a base-3 encoding system: depending on which base was last encoded, a 0, 1 or 2 would correspond with one of the 3 other bases, ensuring the creation of a sequence void of any repetition.

Despite the current high cost of writing and reading DNA, Christophe Dessimoz, another member of the research team, remains hopeful as to the future use of data storage in DNA. “Our analysis shows that for small quantities of data, it is already economically viable for very long term (1,000 years or more), which is relevant for applications such as storing the location of nuclear sites,” says Dessimoz, who is now based atUniversity College London, UK. “If the current pace of technological development continues, within the next decade DNA-based storage will become economical for applications with time horizons of 50 years or more.”

Via Integrated DNA Technologies
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World’s first $1,000 genome enables ‘factory’ scale sequencing for population and disease studies

World’s first $1,000 genome enables ‘factory’ scale sequencing for population and disease studies | Amazing Science |

Illumina, Inc. announced Tuesday that its new HiSeq X Ten Sequencing System has broken the “sound barrier” of human genomics by enabling the $1,000 genome. “This platform includes dramatic technology breakthroughs that enable researchers to undertake studies of unprecedented scale by providing the throughput to sequence tens of thousands of human whole genomes in a single year in a single lab,” Illumina stated.

Initial customers for the HiSeq X Ten System, which will ship in Q1 2014, include Macrogen, based in Seoul, South Korea and its CLIA laboratory in Rockville, Maryland, the Broad Institute in Cambridge, Massachusetts, and the Garvan Institute of Medical Research in Sydney, Australia.

“For the first time, it looks like it will be possible to deliver the $1,000 genome, which is tremendously exciting,” said Eric Lander, founding director of the Broad Institute and a professor of biology at MIT. “The HiSeq X Ten should give us the ability to analyze complete genomic information from huge sample populations. Over the next few years, we have an opportunity to learn as much about the genetics of human disease as we have learned in the history of medicine.”

“The HiSeq X Ten is an ideal platform for scientists and institutions focused on the discovery of genotypic variation to enable a deeper understanding of human biology and genetic disease,” Illumina stated. “It can sequence tens of thousands of samples annually with high-quality, high-coverage sequencing, delivering a comprehensive catalog of human variation within and outside coding regions.”

HiSeq X Ten utilizes a number of advanced design features to generate massive throughput. Patterned flow cells, which contain billions of nanowells at fixed locations, combined with a new clustering chemistry deliver a significant increase in data density (6 billion clusters per run). Using state-of-the art optics and faster chemistry, HiSeq X Ten can process sequencing flow cells more quickly than ever before — generating a 10x increase in daily throughput when compared to current HiSeq 2500 performance.

The HiSeq X Ten is sold as a set of 10 or more ultra-high throughput sequencing systems, each generating up to 1.8 terabases (Tb) of sequencing data in less than three days or up to 600 gigabases (Gb) per day, per system, providing the throughput to sequence tens of thousands of high-quality, high-coverage genomes per year.

Illumina Introduces the HiSeq X™ Ten Sequencing System

Nature: Is the $1000 Genome for real?

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Molecular counting of single molecules using fluorescent proteins

Molecular counting of single molecules using fluorescent proteins | Amazing Science |

To know how many proteins assemble together at the nanoscale is fundamental for understanding protein function. Sometimes, proteins must be in an "oligomeric" state to be functional, although "oligomerization" of certain proteins can also lead to diseases. The ability to determine protein stoichiometry and monitor changes in the balance between monomeric, dimeric and multi-meric proteins can allow scientists to see the differences between a properly functioning cell and a diseased cell. Therefore, there is a great interest in being able to count proteins and determine their stoichiometry.

In a recent study carried out at ICFO, the Institute of Photonic Sciences, the research group of Advanced fluorescence imaging and biophysics, led by Nest Fellow Dr. Melike Lakadamyali was able to quantify the photoactivation efficiency of all the known "ir-reversibly photoswitching fluorescent proteins" and establish a proper detailed reference framework for determining protein stoichiometry. To do this, they used a nanotemplate of known stoichiometry (the human Glycine receptor expressed in Xenopus oocytes) and studied several fluorescent proteins to see the percentage of proteins that was photoactivated. The results of this study have recently been published in Nature Methods.

"Molecular counting" is becoming a closer reality thanks to the discovery of photoactivatable fluorescent proteins and the development of super resolution microscopy. Photoactivatable flourescent proteins change their fluorescence property from dark to bright when exposed to light (i.e. laser light). Through the use of localization-based super-resolution microscopy, researchers are able to photoactivate, image and follow these genetically encoded fluorescent proteins, one at a time, to study what is happening inside a cell at the molecular level. However, despite the "molecular counting" ability that seems intrinsic to the imaging strategy (activating one fluorescent protein at a time should also allow counting how many total fluorescent proteins exist) relating the number of counted fluorescent proteins to actual protein stoichiometry has been difficult. One important reason for this difficult task in counting has been the fact that, until recently, it was not known whether all fluorescent proteins become bright when exposed to laser light. Failure to photoactivate would lead to undercounting since a fraction of probes would be dark and never appear in the image.

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DNA nanorobot from Wyss could potentially seek out cancer cells and cause them to self-destruct

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We've seen various experimental approaches that aim to increase the efficacy of chemotherapy while also reducing its damaging side effects by specifically targeting cancer cells. The latest encouraging development comes from Harvard's Wyss Institute for Biologically Inspired Engineering where researchers have created a barrel-like robotic device made from DNA that could carry molecular instructions into specific cells and tell them to self-destruct. Because the DNA-based device could be programmed to target a variety of cells, it could be used to treat a range of diseases in addition to providing hope in the fight against cancer.

The team based their programmable nanotherapeutic approach on the body's own immune system in which white blood cells circulate in the blood ready to attack an infection where it has developed. Just like white blood cells that are able to hone in on specific cells in distress and bind to them, the researchers created a DNA barrel that can recognize and seek out combinations of cell-surface proteins, including disease markers.

By folding strands of DNA in what is known as the "DNA origami" method, the researchers create a three-dimensional open barrel shape whose two halves are connected by a hinge. The container is held shut by special DNA latches that reconfigure when they find their specific target - cancer cells, for example - causing the two halves to swing open and expose the container's payload. These payloads can be of various types, including molecules with encoded instructions that can interact with surface signaling receptors.

Shawn Douglas, Ph.D., and Ido Bachelet, Ph.D., used the DNA barrel to deliver instructions encoded in antibody fragments to two different types of cancer cells - leukemia and lymphoma. Since leukemia and lymphoma speak different languages the messages were written in different antibody combinations. But the message was the same - activate the cell's so called "suicide gene," which will cause a cell to kill itself through apoptosis.

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Gene therapy method targets tumor blood vessels specifically

Gene therapy method targets tumor blood vessels specifically | Amazing Science |

Working in mice, researchers at Washington University School of Medicinein St. Louis report developing a gene delivery method long sought in the field of gene therapy: a deactivated virus carrying a gene of interest that can be injected into the bloodstream and make its way to the right cells.

In this early proof-of-concept study, the scientists have shown that they can target tumor blood vessels in mice without affecting healthy tissues.

“Most current gene therapies in humans involve taking cells out of the body, modifying them and putting them back in,” said David T. Curiel, MD, PhD, distinguished professor of radiation oncology. “This limits gene therapy to conditions affecting tissues like the blood or bone marrow that can be removed, treated and returned to the patient. Today, even after 30 years of research, we can’t inject a viral vector to deliver a gene and have it go to the right place.”

But now, investigators at Siteman Cancer Center at Barnes-Jewish Hospital and Washington University School of Medicine say they have designed a “targetable injectable vector” – a deactivated virus that homes in on the inner lining of tumor blood vessels and does not get stuck in the liver, a problem that has plagued past attempts.  The findings are reported Dec. 23 in PLOS ONE

“We don’t want to kill tumor vessels,” said senior author Jeffrey M. Arbeit, MD, professor of urologic surgery and of cell biology and physiology. “We want to hijack them and turn them into factories for producing molecules that alter the tumor microenvironment so that it no longer nurtures the tumor. This could stop the tumor growth itself or cooperate with standard chemotherapy and radiation to make them more effective. One advantage of this strategy is that it could be applied to nearly all of the most common cancers affecting patients.”

In theory, Arbeit pointed out, this approach could be applied to diseases other than cancer in which the blood vessels are abnormal, including conditions like Alzheimer’s disease, multiple sclerosis or heart failure.

The viral vector Curiel, Arbeit and their colleagues developed contains a section of DNA called ROBO4 known to be switched on in the cells lining blood vessels within tumors.

In mice, the researchers showed that they could inject the vector into the blood stream and that it accumulated in the tumor vasculature, largely avoiding the lung, kidney, heart and other healthy organs.

Ashlyn Johnson's curator insight, January 7, 2014 1:07 PM

I like this blog because I want to be an crime lab bio technician and you need to know a lot about cells and DNA, and this is availble at the University of Washington.

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Truncated guide RNAs drastically improve specificity of CRISPR-Cas nucleases

Truncated guide RNAs drastically improve specificity of CRISPR-Cas nucleases | Amazing Science |
A simple adjustment to a powerful gene-editing tool may be able to improve its specificity. Investigators have found that adjusting the length of the the guide RNA component of the synthetic enzymes called CRISPR-Cas RNA-guided nucleases can substantially reduce the occurrence of off-target DNA mutations.

Clustered, regularly interspaced, short palindromic repeat (CRISPR) RNA-guided nucleases (RGNs) are highly efficient genome editing tools123. CRISPR-associated 9 (Cas9) RGNs are directed to genomic loci by guide RNAs (gRNAs) containing 20 nucleotides that are complementary to a target DNA sequence. However, RGNs can induce mutations at sites that differ by as many as five nucleotides from the intended target456. A research team recently reports that truncated gRNAs, with shorter regions of target complementarity <20 nucleotides in length, can decrease undesired mutagenesis at some off-target sites by 5,000-fold or more without sacrificing on-target genome editing efficiencies. In addition, use of truncated gRNAs can further reduce off-target effects induced by pairs of Cas9 variants that nick DNA (paired nickases). These results delineate a simple, effective strategy to improve the specificities of Cas9 nucleases or paired nickases.

"Simply by shortening the length of the gRNA targeting region, we saw reductions in the frequencies of unwanted mutations at all of the previously known off-target sites we examined," says J. Keith Joung, MD, PhD, associate chief for Research in the MGH Department of Pathology and senior author of the report. "Some sites showed decreases in mutation frequency of 5,000-fold or more, compared with full length gRNAs, and importantly these truncated gRNAs -- which we call tru-gRNAs -- are just as efficient as full-length gRNAs at reaching their intended target DNA segments."

CRISPR-Cas RGNs combine a gene-cutting enzyme called Cas9 with a short RNA segment and are used to induce breaks in a complementary DNA segment in order to introduce genetic changes. Last year Joung's team reported finding that, in human cells, CRISPR-Cas RGNs could also cause mutations in DNA sequences with differences of up to five nucleotides from the target, which could seriously limit the proteins' clinical usefulness. The team followed up those findings by investigating a hypothesis that could seem counterintuitive, that shortening the gRNA segment might reduce off-target mutations.

"Some of our experiments from last year suggested that one could mismatch a few nucleotides at one end of the gRNA complementarity region without affecting the targeting activity," Joung explains. "That led us to wonder whether removing these nucleotides could make the system more sensitive to mismatches in the remaining sequence."

Based on a natural system a species of bacteria uses against other pathogens, the CRISPR-Cas RGNs most widely used by researchers includes a 20-nucleotide targeting region within the gRNA. To test their theory, the MGH team constructed RGNs with progressively shorter gRNAs and found that, while gRNAs with targeting segments of 17 or 18 nucleotides were as or more efficient than full-length gRNAs in reaching their targets, those with 15- or 16-nucleotide targeting segments had reduced or no targeting activity. Subsequent experiments found that 17-nucleotide truncated RGNs efficiently induced the desired mutations in human cells with greatly reduced or undetectable off-target effects, even at sites with only one or two mismatches.

"While we don't fully understand the mechanism by which tru-gRNAs reduce off-target effects, our hypothesis is that the original system might have more energy than it needs, enabling it to cleave even imperfectly matched sites," says Joung, who is an associate professor of Pathology at Harvard Medical School. "By shortening the gRNA, we may reduce the energy to a level just sufficient for on-target activity, making the nuclease less able to cleave off-target sites. But more work is needed to define exactly why tru-gRNAs have reduced off-target effects."

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First monkeys with customized mutations born, promising better models for human genetic diseases

First monkeys with customized mutations born, promising better models for human genetic diseases | Amazing Science |

Twin cynomolgus monkeys born in China are the first with mutations in specific target genes. This is an important milestone for targeted gene-editing technology, which in turn promises better models for human diseases.

The ultimate potential of precision gene-editing techniques is beginning to be realized. Today, researchers in China report the first monkeys engineered with targeted mutations1, an achievement that could be a stepping stone to making more realistic research models of human diseases.

Xingxu Huang, a geneticist at the Model Animal Research Center of Nanjing University in China, and his colleagues successfully engineered twin cynomolgus monkeys (Macaca fascicularis) with two targeted mutations using the CRISPR/Cas9 system — a technology that has taken the field of genetic engineering by storm in the past year. Researchers have leveraged the technique to disrupt genes in mice and rats23, but until now none had succeeded in primates.

Transgenic mice have long dominated as models for human diseases, in part because scientists have honed a gene-editing method for the animals that uses homologous recombination — rare, spontaneous DNA-swapping events — to introduce mutations. The strategy works because mice reproduce quickly and in large numbers, but the low rates of homologous recombination make such a method unfeasible in creatures such as monkeys, which reproduce slowly.

"We need some non-human primate models," says Hideyuki Okano, a stem-cell biologist at Keio University in Tokyo. Human neuropsychiatric disorders can be particularly difficult to replicate in the simple nervous systems of mice, he says.

Stem-cell researcher Rudolf Jaenisch of the Whitehead Institute for Biomedical Research in Cambridge, Massachusetts, calls the result an interesting demonstration, but says that it offers little scientific insight. "The next step is to see if we can learn anything from it," says Jaenisch, who pioneered the use of transgenic mice in the 1970s.

The combined mutations in Ppar-γ and Rag1 do not represent a particular disease syndrome, says Huang, although each gene is associated with human disorders.The group has yet to fully analyze the monkeys' condition, and must run further tests to assess whether the mutations occurred in all of the animals' cells."Our first aim was to get it done, to get it to work," Huang says. But the finding suggests that researchers could one day model other human conditions involving multiple mutations.

The race is already on to create more CRISPR-modified monkeys, and with greater reliability. Zhang and his colleagues are working to optimize the technology for primate cells, in order to boost mutation efficiency. Okano's team is analyzing unpublished results from monkey models of autism and immune dysfunction, recently created with older gene-editing technologies; they, too, are now trying their luck with CRISPR. And Huang's group is expecting results from eight other pending pregnancies. "There are a lot more things to do," says Huang.

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Single-base genome-editing of human iPS cells without need for antibiotic selection

Single-base genome-editing of human iPS cells without need for antibiotic selection | Amazing Science |

Led by Gladstone Investigator Bruce Conklin, MD, the research team describes in the latest issue of Nature Methods how they have solved one of science and medicine's most pressing problems: how to efficiently and accurately capture rare genetic mutations that cause disease -- as well as how to fix them. This pioneering technique highlights the type of out-of-the-box thinking that is often critical for scientific success.

"Advances in human genetics have led to the discovery of hundreds of genetic changes linked to disease, but until now we've lacked an efficient means of studying them," explained Dr. Conklin. "To meet this challenge, we must have the capability to engineer the human genome, one letter at a time, with tools that are efficient, robust and accurate. And the method that we outline in our study does just that."

One of the major challenges preventing researchers from efficiently generating and studying these genetic diseases is that they can exist at frequencies as low as 1%, making the task of finding and studying them labor-intensive.

"For our method to work, we needed to find a way to efficiently identify a single mutation among hundreds of normal, healthy cells," explained Gladstone Research Scientist Yuichiro Miyaoka, PhD, the paper's lead author. "So we designed a special fluorescent probe that would distinguish the mutated sequence from the original sequences. We were then able to sort through both sets of sequences and detect mutant cells -- even when they made up as little one in every thousand cells. This is a level of sensitivity more than one hundred times greater than traditional methods."

The team then applied these new methods to induced pluripotent stem cells, or iPS cells. These cells, derived from the skin cells of human patients, have the same genetic makeup -- including any potential disease-causing mutations -- as the patient. In this case, the research team first used a highly advanced gene-editing technique called TALENs to introduce a specific mutation into the genome. Some gene-editing techniques, while effective at modifying the genetic code, involve the use of genetic markers that then leave a 'scar' on the newly edited genome. These scars can then affect subsequent generations of cells, complicating future analysis.

Although TALENs, and other similarly advanced tools, are able to make a clean, scarless single letter edits, these edits are very rare, so that new technique from the Conklin lab is needed.

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Human embryonic stem cells can be induced to spontaneously organize into multiple layers of neuroepithelial tissue

Human embryonic stem cells can be induced to spontaneously organize into multiple layers of neuroepithelial tissue | Amazing Science |

During development, the nervous system forms as a flat sheet called the neuroepithelium on the outer layer of the embryo. This sheet eventually folds in on itself to form a neural tube that gives rise to the brain and spinal cord—a process that involves the proliferation and migration of immature nerve cells to form the brain at one end and the spinal cord at the other. Yoshiki Sasai, Taisuke Kadoshima and colleagues from the RIKEN Center for Developmental Biology have now shown that human embryonic stem (ES) cells can spontaneously organize into the cerebral cortical tissue that forms at the front, or ‘brain’ end, of the developing neural tube[1]

Sasai and his colleagues previously developed a novel cell culture technique that involves growing ES cells in suspension, and have shown that these cells can self-organize into complex three-dimensional structures. They have already used this method to grow pieces of cerebral cortex and embryonic eyes from mouse ES cells. And more recently, they have shown that human ES cells can also organize into embryonic eyes containing retinal tissue and light-sensitive cells.

In their most recent work, Sasai’s team treated human ES cells grown using their cell culture system with signaling molecules that induce the formation of nervous tissue from the outer embryonic layer. They found that the cells spontaneously organize into neuroepithelial tissue that then folds up to give a multilayered cortex (see above figure).

During human embryonic development, the neural tube thickens at both ends. In particular, the front end thickens dramatically as waves of cells migrate outward to form the layered cerebral cortex and other parts of the brain. An important finding of the team’s is that the front end of the neural tube appears to thicken due to the growth of radial glial fibers, which span the thickness of the tube and guide migrating cells, rather than due to the accumulation of immature cells within the tube, as previously thought. 

The findings also highlight critical differences between the development of the neural tube in mice and humans. While in humans, the inner surface of the neural tube and the intermediate neuroepithelial zone underneath it contain distinct populations of neural progenitors resembling radial glia, the progenitor population in the latter is not present in the developing mouse cortex.

“Efficient generation of cortical tissues could provide a valuable resource of functional neurons and tissues for medical applications,” says Kadoshima. “By combining this method with disease-specific human induced pluripotent stem cells, it will also be possible to reproduce complex human disorders.”

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Tiny Sponges Seal A Gunshot Wound In 15 Seconds

Tiny Sponges Seal A Gunshot Wound In 15 Seconds | Amazing Science |
An Oregon startup has developed a pocket-size device that uses tiny sponges to stop bleeding fast.

When a soldier is shot on the battlefield, the emergency treatment can seem as brutal as the injury itself. A medic must pack gauze directly into the wound cavity, sometimes as deep as 5 inches into the body, to stop bleeding from an artery. It’s an agonizing process that doesn't always work--if bleeding hasn't stopped after three minutes of applying direct pressure, the medic must pull out all the gauze and start over again. It’s so painful, “you take the guy’s gun away first,” says former U.S. Army Special Operations medic John Steinbaugh.

Even with this emergency treatment, many soldiers still bleed to death; hemorrhage is a leading cause of death on the battlefield. "Gauze bandages just don't work for anything serious," says Steinbaugh, who tended to injured soldiers during more than a dozen deployments to Iraq and Afghanistan. When Steinbaugh retired in April 2012 after a head injury, he joined an Oregon-based startup called RevMedx, a small group of veterans, scientists, and engineers who were working on a better way to stop bleeding.

RevMedx recently asked the FDA to approve a pocket-size invention: a modified syringe that injects specially coated sponges into wounds. Called XStat, the device could boost survival and spare injured soldiers from additional pain by plugging wounds faster and more efficiently than gauze.

The team’s early efforts were inspired by Fix-a-Flat foam for repairing tires. “That’s what we pictured as the perfect solution: something you could spray in, it would expand, and bleeding stops,” says Steinbaugh. “But we found that blood pressure is so high, blood would wash the foam right out.”

So the team tried a new idea: sponges. They bought some ordinary sponges from a hardware store and cut them into 1-centimeter circles, a size and shape they chose on a whim but later would discover were ideal for filling wounds. Then, they injected the bits of sponge into an animal injury. “The bleeding stopped,” says Steinbaugh. “Our eyes lit up. We knew we were onto something.” After seeing early prototypes, the U.S. Army gave the team $5 million to develop a finished product.

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Toddler, an early developmental protein signal hidden amid long 'noncoding' RNA

Toddler, an early developmental protein signal hidden amid long 'noncoding' RNA | Amazing Science |
Scientists at Harvard have identified a previously unknown embryonic signal, dubbed Toddler, that instructs cells to move and reorganize themselves, through a process known as gastrulation, into three layers.

Over the last decade, high-throughput sequencing has revealed a plethora of RNA transcripts that do not seem to encode proteins. These long non-coding RNAs (lncRNAs) have emerged as key players in gene regulation. In the past, annotation of lncRNAs has largely relied on the computational identification of transcripts that lack coding features. The recent development of ribosome profiling has provided a high-throughput method of assessing ribosome engagement over the whole transcriptome at near-nucleotide resolution.

Harvard scientists now integrated ribosome profiling and RNA-Seq data from a zebrafish developmental time course using a random forest machine learning approach to distinguish different modes of translation. They first showed that it is possible to discriminate ribosome engagement over transcript leaders and transcript trailers from protein coding open reading frames (ORFs). Using this result they classified lncRNAs into the category that they resembled the most (protein-coding, leader-like, trailer-like). This surprisingly revealed that many lncRNAs were engaged by ribosomes in a similar manner to upstream ORFs (uORFs) in transcript leaders. Unlike canonical proteins, these lncRNAs do not seem to have a single dominant ORF that is translated instead they often contain multiple short ORFs with more dispersed translation.

Several observations suggest that these translated lncRNA-ORFs might generate proteins that are likely to be non-functional. First, their diminutive size (sometimes just a single amino acid) is often incompatible with a functional peptide. Second, they show a distinct lack of conservation and in particular conservation at the amino acid level. Instead, ribosomal engagement of this subclass of lncRNAs may have a regulatory role. Translation can affect the stability of transcripts and/or their localization through a number of pathways like nonsense-mediated RNA decay (NMD) that the cell could utilize to modulate the lncRNA expression levels. Alternatively, spurious translation could potentially be functional over evolutionary time as a source of new proteins. These data also revealed that computational classification should be used with care as some pipelines can result in up to 45% of false positives (proteins). The researchers show that using a combination of homology and evolutionary measures results in a good classification and should be adopted as the standard in the field.

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Cheaper, faster and safer way of converting cells to stem cells, using acid shock

Cheaper, faster and safer way of converting cells to stem cells, using acid shock | Amazing Science |

Scientists in Japan showed stem cells can now be made quickly just by dipping blood cells into acid. The latest development, published in the journal Nature, could make the technology cheaper, faster and safer.

The researchers report a unique cellular reprogramming phenomenon, called stimulus-triggered acquisition of pluripotency (STAP), which requires neither nuclear transfer nor the introduction of transcription factors. In STAP, strong external stimuli such as a transient low-pH stressor reprogrammed mammalian somatic cells, resulting in the generation of pluripotent cells. Through real-time imaging of STAP cells derived from purified lymphocytes, as well as gene rearrangement analysis, the scientists found that committed somatic cells give rise to STAP cells by reprogramming rather than selection. STAP cells showed a substantial decrease in DNA methylation in the regulatory regions of pluripotency marker genes. Blastocyst injection showed that STAP cells efficiently contribute to chimaeric embryos and to offspring via germline transmission. They could also demonstrate the derivation of robustly expandable pluripotent cell lines from the obtained STAP cells. Thus, these findings indicate that epigenetic fate determination of mammalian cells can be markedly converted in a context-dependent manner by strong environmental cues.

The finding has been described as "remarkable" by the Medical Research Council's Prof Robin Lovell-Badge and as "a major scientific discovery" by Dr Dusko Ilic, a reader in stem cell science at Kings College London.

Dr Ilic added: "The approach is indeed revolutionary.

"It will make a fundamental change in how scientists perceive the interplay of environment and genome." But he added: "It does not bring stem cell-based therapy closer. We will need to use the same precautions for the cells generated in this way as for the cells isolated from embryos or reprogrammed with a standard method."

And Prof Lovell-Badge said: "It is going to be a while before the nature of these cells are understood, and whether they might prove to be useful for developing therapies, but the really intriguing thing to discover will be the mechanism underlying how a low pH shock triggers reprogramming - and why it does not happen when we eat lemon or vinegar or drink cola?"

Scientist accused of falsifying data

Dr. Stefan Gruenwald's insight:

This story is under investigation. It was published in Nature but the data seem to be partially / completely false.

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Remote-controlled spermbots could be used to fertilize eggs

Remote-controlled spermbots could be used to fertilize eggs | Amazing Science |

Researchers have hijacked sperm cells to create spermbots that can be remotely controlled through magnetic fields, to go to the desired location.

Hijacking sperm cells to create little robots might seem far out, but that's exactly what researchers from the Dresden Institute for Integrative Nanosciences have done. Their "spermbots" consist of live sperm cells in little tubes, that can be magnetically controlled to move in a desired direction until they reach their destination and do their job – they're currently robust enough to even guide a specific sperm cell to an egg cell. The scientists hope that further development will allow the technology to offer a viable alternative to parents trying to have a child through in-vitro fertilization. When perfected, the spermbots could also be used as a safe means for drug delivery and gene manipulation.

One of the major challenges in creating micro robots that can potentially travel within the human body is the issue of a safe fuel source. Nanobots with engines efficient enough to propel themselves through bodily fluids need to carry fuel that's often toxic to the human body, and sometimes these machines can pass through into the cells and affect their functioning. To overcome these problems, the Dresden team began looking at safer alternatives to artificial nano engines.

"We thought of using a powerful biological motor to do the job instead and we came up with the flagella of a sperm cell, which is physiologically less problematic," Professor Oliver G. Schmidt, the Institute's Director, tells Gizmag. "The idea came to us five years ago when I noticed that sperm cells are of similar size to microtubes we can fabricate."

To create biorobots out of sperm cells, the researchers began working with bovine (bull) sperm cells – which are similar in size to human sperm cells. The first step was to create thin conical magnetic tubes capable of trapping sperm cells out of a titanium and iron film. The microtubes are rolled up in a way that makes one end larger than the other, with a diameter that's slightly larger than that of a bull sperm head.

Via Jeff Morris
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Gene therapy could be used to treat blindness: Sight restored to partially blind patients

Gene therapy could be used to treat blindness: Sight restored to partially blind patients | Amazing Science |

Surgeons in Oxford have used a gene therapy technique to improve the vision of six patients who would otherwise have gone blind. The operation involved inserting a gene into the eye, a treatment that revived light-detecting cells. The doctors involved believe that the treatment could in time be used to treat common forms of blindness. Prof. Robert MacLaren, the surgeon who led the research, said he was "absolutely delighted" at the outcome.

Mr. Wyatt has a genetic condition known as choroideremia, which results in the light-detecting cells at the back of the eye gradually dying. He was still just about able to see when he had the operation. His hope was that the procedure would stop further deterioration and save what little sight he had left.

He, like another patient in Prof. MacLaren's trial, found that not only did the operation stabilise his vision - it improved it. The other subjects, who were at earlier stages in their vision, experienced improvements in their ability to see at night. Mr. Wyatt is now able to read three lines further down in an optician's sight chart. "I felt that I had come to the edge of an abyss," he said. "I looked down at total blackness. Prof. MacLaren tapped me on the shoulder and said 'come this way, it's possible to see again'."

Another of the patients who underwent the treatment, Wayne Thompson, said he had noticed an immediate effect after the operation. "My color vision improved. Trees and flowers seemed much more vivid and I was able to see stars for the first time since I was 17 when my vision began to deteriorate," he explains. Mr. Thomson said he had spent his life resigned to the fact that he would go blind. "I've lived the last 25 years with the certainty that I am going to go blind and now (after the operation) there is the possibility that I will hang on to my sight," he said.

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Stanford bioengineers redesign protein motors to create novel nanomachines

Stanford bioengineers redesign protein motors to create novel nanomachines | Amazing Science |

Stanford scientists genetically engineer versions of myosin proteins that transport biological materials in cells to illuminate design features that keep these protein motors on track. Inside our cells, proteins known as myosins can act as a delivery service for biological materials. To better understand how molecular motors move, Stanford bioengineers have built experimental versions of the proteins, changing the way these transporters get around. Led by Zev Bryant, an assistant professor of bioengineering at Stanford, a team of researchers has genetically engineered “mutant” myosins with new features such as gearshifts and improved traction. The group’s most recent findings are published in the January issue of Nature Nanotechnology, where they are highlighted alongside other studies of molecular motors.

“You look at biology, and you see motors that have diverse mechanical properties, and you want to understand how these arise,” Bryant said. “You test your understanding by trying to build something new.” Molecular motors are a class of proteins that make up the moving machinery of cells. Myosins are one family of molecular motors. Some of them can shuttle biomolecules from one region of the cell to another.

These myosins move along microscopic filaments made of the protein known as actin. These actin filaments are one component of the cytoskeleton, or internal support structure of the cell. Bryant wanted to test his understanding of how evolution has designed these myosin proteins to shuttle cellular freight. Funded by an NIH “New Innovator” Award, members of the group launched a series of experiments in 2008 that steered their myosin research in a new direction. They began engineering myosins with extra parts to give natural myosins new capabilities.

Natural myosins, for example, see actin filaments as one-way tracks. To better understand this one-directional motion, Bryant challenged his group to design mutant myosins that could move forward and backward on command. The researchers engineered myosin motors with extra components that behaved like a molecular gearshift. In a 2012 Nature Nanotechnology report, the researchers showed that they could shift their mutant myosin motion between forward and reverse. However, these two-way myosins had trouble hanging onto their actin tracks.

“When we engineer motors to have new capabilities, we often sacrifice some capabilities that they already had,” Bryant said. So the group focused on creating motors that excelled at hanging on.

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Micro Electron Diffraction Could Revolutionize Structural Biology Studies

Micro Electron Diffraction Could Revolutionize Structural Biology Studies | Amazing Science |

For structural biologists, the first step in determining a protein's precise molecular structure is often the hardest: coaxing the protein to grow into the orderly, three-dimensional crystals that are the starting material for most structural studies. For particularly difficult cases, it can take years to generate usable crystals – and sometimes the protein never crystallizes despite intense effort. Howard Hughes Medical Institute (HHMI) scientists have developed a new method that generates a high-resolution protein structure from crystals one-million times smaller than those needed for X-ray crystallography, the most common method for determining protein structure.

The new technique, called MicroED (micro electron diffraction), has the potential to accelerate structural biologists' efforts and to expand the repertoire of proteins whose high-resolution structures can be solved. “Biochemically, it is always easier to generate smaller crystals,” says structural biologist Tamir Gonen, who developed the technique with colleagues in his lab at HHMI's Janelia Farm Research Campus. “There are many proteins where you either don't get crystals or you get crystals that are very, very small. They might be good enough for microED.”

Gonen and colleagues describe MicroED in a report published November 19, 2013, in the open access journal eLife. Using Gonen’s new technique, scientists can use electrons from an electron microscope to determine the structure of protein microcrystals. This approach, known as electron crystallography, had previously been limited to studies of proteins that could be grown into very thin, two-dimensional crystals. “People have put 3D crystals in electron microscopes before, but no one was able to solve their structures,” Gonen says.

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NATURE: The Method of the Year for 2013 is… single-cell sequencing

NATURE: The Method of the Year for 2013 is… single-cell sequencing | Amazing Science |

Single-cell sequencing edged out other contenders as our choice of Method of the Year in 2013. These techniques really came into their own in 2013 and are fast providing new insights into the workings of single cells that ensemble methods are incapable of.

Back in 2008 we chose next-generation sequencing as our Method of the Year not only because of how the new techniques would improve performance in conventional sequencing applications, but also because they opened up whole new applications, unthinkable with traditional Sanger sequencing. Our choice of Method of the Year in 2013 bears this out, as none of these single-cell sequencing applications would be possible without next-generation sequencing. And in some applications the sequencing is used almost exclusively for identifying and counting tagged molecules.

Our choice likely comes as a surprise to all those who were certain that we would pick CRISPR/Cas9 technology for targeted genome modification. This is certainly an exciting technology, and not only for genome engineering, but also for epigenome editing as described in a Method to Watch. But genome editing with engineered nucleases was our pick for the 2011 Method of the Year and although CRISPR/Cas9 provides a huge practical improvement by largely dispensing with the need to engineer the nuclease and relying instead on a programmable guide RNA, the advance over 2011 is mostly one of ease-of-use.

Methods to investigate biology at the level of single cells have been of keen interest to Nature Methods since the journal started. Our first research article from Robert Singer described a paraffin-embedded tissue FISH (peT-FISH) method to simultaneously detect expression of several genes in situ in single cells while maintaining tissue morphology (Capodieci, P. 2005). This was followed by many other imaging-based methods for such things as measuring cell growth (Groisman, A. 2006), quantifying mRNA (Raj, A. 2008) and protein (Gordon, A. 2006) levels, profiling intracellular signaling (Krutzik, P.O. & Nolan, G.P. 2006) (Loo, L.-H. 2007) and DNA insertion-site analysis (Schmidt, M. 2008) in single cells.

The publication of M. Azim Surani’s article on mRNA-Seq whole-transcriptome analysis of a single cell (Tang, F. 2009) in 2009 helped signal the rise of sequencing-based methods for single-cell analysis. But even two years later the Reviews and Perspectives in our supplement on single-cell analysis were more focused on imaging-based than sequencing-based aproaches to single-cell analysis.

It was only in 2013 that we finally saw an explosion of original research articles using or reporting single-cell sequencing methods in Nature-family journals. Numerous studies reported new biological results that relied on sequencing of whole or partial genomes or transcriptomes from single cells.

Nature's Method of the Year special feature has three Commentaries by researchers in the field, including some of the earliest developers and users of methods for single-cell analysis. An Editorial, News Feature and Primer describe our choice and provide helpful background information. We hope you enjoy the selection of articles in our special feature.

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