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The genome editor CRISPR cuts DNA with help from a guide RNA (green and red) and a Cas9 enzyme (outline) that latches onto a three-base sequence (yellow).


Upgrade makes genome editor CRISPR more muscular, precise

You wouldn’t know it from the excitement generated by the revolutionary genome editing method known as CRISPR, but as practiced now, it is far from perfect. Its standard components can find and cut DNA in only a limited fraction of the genome, and its molecular scissors are wobbly, leading to “off-target” mutations. Many groups are trying to do better, and now, a team led by chemist David Liu at Harvard University has engineered a version of CRISPR that potentially is both more dexterous and more precise.

“This is very impressive and important work,” says CRISPR pioneer Erik Sontheimer of the University of Massachusetts Medical School in Worcester.

CRISPR comes in many flavors, but they all depend on a guide molecule composed of RNA to carry a DNA-cutting enzyme—the most commonly used one is known by the shorthand Cas9—to a specific stretch of the genome. This complex, however, homes in on DNA landing pads that have specific molecular features. The enzyme in the standard CRISPR toolkit, called spCas9 for its natural source, the bacterium Streptococcus pyogenes, can only land on genome segments that have at one end a specific three-base trio: N, where N is any of DNA’s four bases, followed by two guanines (Gs). Only about one-sixteenth of the 3.2-billion-base human genome has the right sequence. “That’s been a real limitation,” Liu says.

The new work, reported online in the 28 February issue of Nature, modifies the Cas9 enzyme, creating at least four times as many potential docking sites. In theory, this could allow researchers to, say, cripple or replace many parts of genes associated with human disease that CRISPR currently cannot touch.

Liu’s lab began by engineering a large variety of slightly altered spCas9s. The group then selected for ones that could use a broader range of the 64 possible, three-base landing pads—technically referred to as protospacer adjacent motifs, or PAMs. They’ve dubbed their new enzymes xCas9s, and the best one works with NGN, a sequence that occurs in one-fourth of the genome.

Liu expected that in return for gaining the ability to latch onto more places, xCas9 would pay a penalty: more of the potentially dangerous off-target cuts that concern researchers hoping to unleash CRISPR in medicine. After all, conventional thinking holds that Cas9, naturally part of a bacterial immune strategy, evolved to be as promiscuous in its DNA binding as it could be without compromising specificity. “PAM binding is supposed to be the gatekeeper, and if your gatekeeper is drunk and lets lots of Cas9 into the dance, that should screw up the targeting,” Liu explains. But the opposite happened. “If you ask me for a detailed mechanistic explanation for why that is, my answer is, ‘I don’t know,’” he says.

Stanley Qi, a CRISPR researcher at Stanford University in Palo Alto, California, says this win-win situation is “amazing,” and should excite many labs. “The real test here is if people rush to use this xCas9 while forgetting about the original version,” Qi says. “At least in my lab, we are very eager to try this out for our applications.”

Liu cautions that the standard Cas9 has proved itself over the years; his lab has only tested the new xCas9 on a few dozen sites in the genome so far, compared with the thousands the original has been shown to hit. “I’m not 100% sure xCas9 is going to be flat out better than spCas9,” Liu says. “I want everyone to test it because I want to know the answer.”


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Science Daily

 Linking virus sensing with gene expression, a plant immune system course-corrects
March 6, 2018
American Society for Biochemistry and Molecular Biology

Plant immune systems, like those of humans and animals, face a difficult balancing act: they must mount responses against ever-evolving pathogens, but they must not overdo it. Immune responses require energy and resources and often involve plants killing their own infected cells to prevent the pathogens from spreading.

Researchers at Durham University in the UK have identified a crucial link in the process of how plants regulate their antiviral responses. The research is published in the March 2 issue of the Journal of Biological Chemistry.

Martin Cann’s lab at Durham, in collaboration with the laboratories of Aska Goverse at Wageningen University and Frank Takken at the University of Amsterdam, studied a receptor protein called Rx1, which is found in potato plants and detects infection by a virus called potato virus X.

Binding to a protein from the virus activates Rx1 and starts a chain of events that results in the plant mounting an immune response. But the exact sequence of cellular events — and how Rx1 activation was translated into action by the rest of the cell — was unknown.

“Our study revealed an exciting, and unexpected, link between pathogen attack and plant DNA,” Cann said.

Specifically, the study showed that Rx1 joins forces with a protein called Glk1. Glk1 is a transcription factor, meaning it binds to specific regions of DNA and activates genes involved in cell death and other plant immune responses. The team found that when Glk1 bound to virus-activated Rx1, it was able to turn on the appropriate defense genes.

Interestingly, when the viral protein was absent, Rx1 seemed to have the opposite effect — actually keeping Glk1 from binding to DNA. In this way, it prevented an inappropriate immune response.

“The immune response involves reprogramming the entire cell and also often the entire plant,” Cann said. “An important part of this regulatory process is not only allowing activation but also making sure the entire system is switched off in the absence of infection.”

As over a third of the annual potential global crop harvest is lost to pathogens and pests, breeding plants with better immune systems is an important challenge. Understanding how this immune system is regulated at the appropriate level of activity gives the researchers more ideas of points in the immune signaling pathway that could targeted to increase the plant’s baseline ability to resist disease.

“To increase (crop) yield, there is an urgent need for new varieties that are resilient to these stresses,” Cann said. “A mechanistic understanding of how plants resist or overcome pathogen attack is crucial to develop new strategies for crop protection.”

Story Source:

Materials provided by American Society for Biochemistry and Molecular Biology. Note: Content may be edited for style and length.

Journal Reference:

  1. Philip D. Townsend, Christopher H. Dixon, Erik J. Slootweg, Octavina C. A. Sukarta, Ally W. H. Yang, Timothy R. Hughes, Gary J. Sharples, Lars-Olof Pålsson, Frank L. W. Takken, Aska Goverse, Martin J. Cann. The intracellular immune receptor Rx1 regulates the DNA-binding activity of a Golden2-like transcription factor. Journal of Biological Chemistry, 2018; 293 (9): 3218 DOI: 10.1074/jbc.RA117.000485

American Society for Biochemistry and Molecular Biology. “Linking virus sensing with gene expression, a plant immune system course-corrects.” ScienceDaily. ScienceDaily, 6 March 2018. <www.sciencedaily.com/releases/2018/03/180306153726.htm>.

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UGA Today

Science & Technology

UGA researchers develop new method to improve crops

Scientists examining plants

William Jordan (left) and Lexiang Ji look over one of many sets of Arabidopsis thaliana, which were used to research a new plant breeding technique.


Technique using plant’s own DNA could produce crops that are more resistant to drought and disease


A team of University of Georgia researchers has developed a new way to breed plants with better traits. By introducing a human protein into the model plant species Arabidopsis thaliana, researchers found that they could selectively activate silenced genes already present within the plant.

Using this method to increase diversity among plant populations could serve to create varieties that are able to withstand drought or disease in crops or other plant populations, and the researchers have already begun testing the technique on maize, soy and rice.

They published their findings in Nature Communications.

The research project was led by Lexiang Ji, a doctoral student in bioinformatics, and William Jordan, a doctoral student in genetics. The new method they explored, known as epimutagenesis, will make it possible to breed diverse plants in a way that isn’t possible with traditional techniques.

“In the past this has been done with traditional breeding. You take a plant, breed it with another plant that has another characteristic you want to create another plant,” said Jordan. “The problem with that is getting an individual that has all of the characteristics you want and none of the characteristics that you don’t want. It’s kind of difficult. With our new technique, you can modify how the genes are turned on and off in that plant without having to introduce a whole other set of genes from another parent.”

The idea for the method evolved originally from working in the lab with department of genetics professor Robert Schmitz, the corresponding author on the study. In his lab, researchers were studying DNA methylation, which controls expressed genetic traits, and creating maps of where DNA methylation is located in many plant species, including crops. When DNA methylation is removed, researchers found that they could selectively turn on previously silenced genes in the underlying genome of the plant.

“We saw repeatedly that lots of genes are silenced by DNA methylation and thought it was kind of curious,” said Schmitz. “There are lots of discussions you can have about why these exist, but the reality is that they are there. So we wondered, how can we leverage them? Let’s use the plant already in the field and reawaken some of those silenced genes to generate trait variation.”

To turn these dormant or silenced genes on, researchers introduced a human enzyme, known as a ten–eleven translocation enzyme, to plant seedlings using specially modified bacteria as a delivery vector. Introducing this human protein allows researchers to remove DNA methylation and thereby turn on previously silenced genes.

Figuring out the best way to introduce the protein to the plant species has been a trial and error process. With Ji’s expertise in bioinformatics, researchers are able to look at large sets of data about their experiment and make decisions on how to best proceed with the project.

“The data has really helped us brainstorm and coordinate what we should do next,” said Ji. “That was particularly important in the beginning of this project because we just didn’t know what was going to happen with this new technique.”

“Thousands of years ago you’d plant out hundreds of plants and one of them does really well so you’d breed out generations of that plant. Doing this though, you narrow down the genetic diversity until they’re basically very, very similar,” said Jordan. “While that’s beneficial for yield or other plant characteristics that you might want, if there’s a stress that they’re not well adapted to because they’re all so similar they’re all going to respond in the same way. That creates a potentially vulnerable crop.”

“If they don’t have the genetic differences to respond, then it can really wipe out crops,” added Schmitz. “This isn’t a savior, but it’s an alternative strategy that has not been tried before. The idea is to access genes that people haven’t been studying because they’re not expressed but they’re there. We think this method to reactivate these genes could lead to increased trait variation which could be useful for biotechnology applications.”

The study, “TET-mediated epimutagenesis of the Arabidopsis thaliana methylome,” was published in Nature Communications March 1, 2018, and is available at: https://www.nature.com/articles/s41467-018-03289-7

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Fundação de Amparo à Pesquisa do Estado de São Paulo

Public Release: 

Computer models allow farmers to diversify pest management methods

A technology developed by Brazilian researchers can help fighting highly resistant agricultural pests by analyzing the connections between the pests’ patterns of dispersal in crops and different configurations in diversified intercropping systems.


An article published in Scientific Reports shows some results from a study supported by the São Paulo Research Foundation – FAPESP which have developed mathematical models to describe the movements of agricultural pests, aiming at better understanding how these pests disperse in agricultural areas.

“The idea is to use computer models to design strategies capable of reducing the damage done to crops by pest populations and containing their expansion on plantations,” said Wesley Augusto Conde Godoy, coordinator of the project and also a professor at the University of São Paulo’s Luiz de Queiroz College of Agriculture (ESALQ-USP), institution where the project took place. The investigation featured collaboration from researchers from the São Paulo State University (UNESP) in Botucatu, São Paulo State, Brazil.

First, the researchers modeled the movements of the cucurbit beetle, Diabrotica speciosa, which attacks several crops, such as soybeans, corn and cotton.

Using computer modeling, they found that spatial configurations in diversified intercropping systems (growing two or more crops in proximity) could favor or inhibit pest dispersal. “We observed that the presence of corn strips distributed across farmed fields could reduce spatial dispersal of the insects,” Godoy said.

Motivated by the results obtained with D. speciosa, they investigated possible applications of computer modeling to describe the spatial dynamics of other agricultural pests, such as the Fall armyworm (S. frugiperda), an insect which has developed resistance to Bt corn, cotton and soybeans.

In order to delay the development of S. frugiperda and other pests’ resistance to transgenic crops – which have been modified to repel parasites through the inclusion of genetic material from Bacillus thuringiensis Berliner (Bt) bacteria -, technicians have advised farmers to create refuges, strips within Bt crop fields of the same crop without a Bt trait.

Refuges are intended to ensure the maintenance of individuals susceptible to Bt technology within the pest population. They mate with resistant individuals, and this prevents the population as a whole from developing resistance to Bt toxins, Godoy explained. “It’s been demonstrated that the larger the refuge area is, the lower the frequency of Bt-resistant individuals,” he said.

Using a cellular automata-based computer model that predicts the movements of insects, the researchers measured the effectiveness of three different refuge configurations: mixed seeds, random blocks, and strips.

“We succeeded in identifying the best refuge configuration and size to delay the development of Bt crop resistance in S. frugiperda,” Godoy said.

Comparing movement patterns

The researchers combined the computer model with data on the insect’s movements obtained in the laboratory to analyze and compare its behavior on leaves of Bt and non-Bt cotton. The results of the study showed that the insect moved around more on Bt cotton leaves than non-Bt cotton leaves.

“We don’t yet know what mechanisms may trigger this behavior,” Godoy said. “However, the findings so far have important practical implications because they may correlate with faster development of Bt resistance.”

Less movement on non-Bt leaves could be associated with adaptation cost. This is frequently the case for resistant populations of this insect in the absence of selection pressure, he explained.

“We plan to continue investigating this problem, as continuation of the research could produce significant contributions to pest management programs by improving crop configuration to delay the development of resistance to GM crops among these and other insects,” Godoy said.


About São Paulo Research Foundation (FAPESP)

The São Paulo Research Foundation (FAPESP) is a public institution with the mission of supporting scientific research in all fields of knowledge by awarding scholarships, fellowships and grants to investigators linked with higher education and research institutions in the State of São Paulo, Brazil. FAPESP is aware that the very best research can only be done by working with the best researchers internationally. Therefore, it has established partnerships with funding agencies, higher education, private companies, and research organizations in other countries known for the quality of their research and has been encouraging scientists funded by its grants to further develop their international collaboration. For more information: http://www.fapesp.br/en.

Disclaimer: AAAS and EurekAlert! are not responsible for the accuracy of news releases posted to EurekAlert! by contributing institutions or for the use of any information through the EurekAlert system.

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Plague-Resistant Tomatoes Developed through Genetic Engineering

A study conducted by researchers at the Plant Molecular and Cellular Biology Institute (IBMCP), a joint venture of the Universitat Politècnica de València and the Spanish National Research Council (CSIC), reveals how genetically modified tomato plants have increased resistance towards Tuta absoluta insect plagues.

An estimated 40% of the worldwide annual crop production is lost to plagues and pathogens, and 13% to insects. Luis Cañas, researcher of the CSIC at the IBMCP, explains that “the miner insect Tuta absoluta has become one of the main plagues that threaten tomato plantations across the world, and without the appropriate management it can cause losses of between 80% and 100% of their production.”

The researchers turned to genetic engineering to strengthen the tomato plant by giving it defensive genes such as the protease inhibitors in barley. A serine proteinase inhibitor (BTI-CMe) and a cysteine proteinase inhibitor (Hv-CPI2) were investigated, isolated from the barley plant, on the Tuta absoluta insect. Both inhibitors were tested separately, as well as together in transgenic tomato plants. The Tuta absoluta larvae which were fed the double transgenic plants showed noticeable weight loss, and only 56% of the larvae reached their adult stage. Those that reached their adult phase had wing deformities and fertility reduction.

For more details, read the news release at R&I World

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Delta f perss

cotton bollworm

Cotton diseases and insect control as resistance appears

Growers will have to make some difficult decisions related to foliar diseases and Bt-resistant worm pests.

Brad Robb | Jan 25, 2018

Mid-South cotton growers face several tough decisions for 2018 as they deal with hard-to-control diseases and the increasingly difficult problem of Bt-resistant worm pests.

Tucker Miller, president, Miller Entomological Service Inc., Drew, Ms., speaking Thursday at the National Conservation Systems Conferences in Memphis, Tn., said growers will need to look closely at varieties as well as other management options.

Miller, a frequent speaker at the conference, spoke to a packed meeting room about his experience over the last several years with bacterial blight and target leaf spot, potassium-associated foliar diseases. “Growers are going to need to make several important variety selection decisions this coming season, and those decisions need to be made based on good information,” says Miller. “They’ll need a variety that is resistant to bacterial blight and, because there is no variety that provides resistance to target leaf spot, they’ll have to consider other management options to try to control its level and to lessen the effect or impact of the leaf disease.”

Options may include decreasing seeding rates to produce a thinner stand, aggressive Pix management, possibly growing skip row cotton, or selecting a variety or row configuration that lends itself to a more open canopy to help minimize the spread of target leaf spot. “Growers might also try to manage this disease with more timely irrigation methods or even less irrigation,” says Miller. “Leaf shed was so bad in many parts of the fields I worked, if you squatted and looked down the row, you could see a rabbit two-hundred yards away.”

The Mid-South is supposedly in the low to medium risk range of the country for this problem, but Miller questions those range boundaries. Several factors, including irrigation and over-fertilization of nitrogen, may be exacerbating the problem. “It’s hard to get farmers to cut back to 80 or 90 units of nitrogen when they’re accustomed to putting out 120, and they don’t want to run out,” says Miller. “Fungicides are another option, but at $40 an acre, if you spray it twice, I just don’t know if it’s a cost effective application.”


Based on one data set Miller received from 2016 target spot research, a fungicide application to control the disease may provide a significant yield increase only 20 percent of the time. “It’s difficult for me to suggest an application of fungicide at first or second bloom with an 80 percent chance it won’t help,” says Miller.

Miller also talked frankly about the resistant worm (heliothis) problems many growers across the Mid-South and Southeast experienced last year. According to Miller, the problem started with a generation of worms exposed to Bt corn with the two identical proteins found in Bollgard ll or WideStrike ll cotton varieties. “When worms go through a generation and come out of corn then move to cotton, they’re exposed to the same proteins twice, but the second time, they’re surviving,” says Miller.

Dried bloom tags were everywhere when Miller scouted some Bt fields last year. At one point of the season, a report to one of his grower customers listed a high-dollar combination shot of Besiege, Acephate and Pix. “I recommended Pix to control plant growth, Acephate for plant bugs and Besiege for worms on July 16, and by July 29, we had to do it again in one field of Bollgard ll cotton,” says Miller.

One problem researchers across the board are concern about is how long the third protein – VIP—will remain viable if the same scenario presents itself once growers begin planting corn and cotton with the VIP protein. “It’s going to take some careful management for sure,” says Miller.

The 21st Annual National Conservation Systems Conferences will likely set a new record. Growers, Extension specialists, and agricultural researchers covering many disciplines present over 120 presentations over the day-and-a-half conference.


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Uganda: GMO bananas

Plantwise Blog

Is It Time for Genetically Modified Bananas in Uganda?


Uganda is the world’s second largest producer of banana crop, with individuals consuming around 1.5 pounds of banana every day. Due to this major need for the success of banana crops within the country, plant pests and diseases are ever more threatening.

An example of this is with the invasion of banana bacterial wilt in the last decade, which was predicted to destroy 90% of Uganda’s banana crop at a loss of around $4 billion, however the spread was stopped after a series of expensive government interventions. With global warming threatening an increase in plant pest and disease spread on a global scale, it is not surprising that the country is considering the use of genetic modification as a solution to this issue.

Researchers have been undertaking field tests using disease resistant genetically modified (GM) bananas for years, however most countries in Africa ban farmers from commercially growing such crops. Until recently, South Africa was the only country in Sub-Saharan Africa to allow the commercial use of GM food. Uganda was added to the list of pro-GMO countries in October 2017, as parliament passed the National Biosafety Act 2017 which opens up agricultural biotechnology i.e. large scale field testing and the commercial use of GM crops.

Woman with bananas

“Finally, banana farmers will be able to access varieties of banana resistant to bacterial wilt, and the people, especially children, can finally eat bananas and other foods rich in Vitamin A” said Patrick Nanteza who works with the National Banana Programme in Kawanda.

For many, the passing of this bill is a daunting step, as it opens up the unexamined risks of producing and consuming GM crops. For more information in regards to the debate on the use of GMO’s commercially, please see the links below:

Biotechnology is a key component of the solution to the global demand for food and the increasing threats to food security. Specifically within Uganda, roughly 75% of the country is involved in agriculture; however there is a nation-wide food shortage due to recent climatic events. In 2017 alone, over 70% of the country’s arable land was affected by drought and unpredictable rainy seasons.

For further information on this subject, the original article published by PRI is available here:


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