Botanic gardens are also part of the picture. In another paper in the same issue, Yang Li et al. from the Key Laboratory of Tropical Plant Resources and Sustainable Use at Xishuangbanna Tropical Botanical Garden in Kunming (Yunnan) and the University of the Chinese Academy of Sciences in Beijing present research on DELLA-interacting proteins in Arabidopsis. Here the authors show that bHLH48 and bHLH60 are transcription factors involved in GA-mediated control of flowering under long-day conditions.
Naturally, research on rice is important. Wei Jiang et al. from the National Key Laboratory of Crop Genetic Improvement, Huazhong Agricultural University (Wuhan) describe their research on WOX11 and the control of crown root development in the nation’s grain of choice, which will be important for breeders looking to increase crop yields and resilience.
Shenzehn has grown rapidly and is now highly significant for life science as home to the China National GeneBank (CNGB) project led by BGI Genomics. The vision as set out by Huan-Ming Yang, chairman of BGI-Shenzhen, is profound – from sequencing what’s already here, often in numbers per species, to innovative synthetic biology.
Shenzehn is also home to another significant institution, the beautiful and scientifically important Fairy Lake Botanic Garden. At the IBC, the importance of biodiversity conservation for effective, economically focused plant science, but also for so many other reasons to do with our intimate relationship with plants and continued co-existence on the planet, was a central theme.
The research highlighted in Journal of Experimental Botany is part of the wider, positive growth of plant science (and, indeed, botany) not just in China, but worldwide. The Shenzehn Declaration on Plant Sciences with its seven priorities for strategic action, launched at the congress, will be a guide for the right development in coming years.
Could you begin by telling us a little about your research?
I am a plant physiologist specializing in seed biology. I have a long research record on various aspects of seeds, including the mechanisms and regulation of germination and dormancy, desiccation tolerance, as well as issues in seed technology. Being six years from retirement now, I decided to extend my desiccation tolerance studies from seeds to resurrection plants, which display vegetative desiccation tolerance. I strongly believe that unveiling of the mechanism of vegetative desiccation tolerance may help us create crops that are truly tolerant to severe drought, rather than (temporarily) resistant.
How did you become interested in this field of study, and how has your career progressed?
As with many things in life, it was coincidence. I majored in plant biochemistry and applied for a PhD position in seed biology. After obtaining the degree I was offered a tenure track position in seed physiology by the Laboratory of Plant Physiology at Wageningen University, where I still work as a faculty member. My career has progressed nicely and I am an authority in the field of seed science, editor-in-chief of the journal Seed Science Research, and will become the President of the International Society for Seed Science in September of this year.
I see my current work on vegetative desiccation tolerance as a highlight in my professional life. I have always been more interested in the desiccation tolerance of seeds until about five years ago, when my current collaborator Prof Jill Farrant of the University of Cape Town, South-Africa, made me enthusiastic about these wonderful resurrection plants. We started to work together and published our first study recently in Nature Plants.
In your recent paper, you sequenced the genome of the resurrection plant, Xerophyta viscosa, which can survive with less than a 5% relative water content. How is it possible for a plant to lose so much of its water and still survive?
These plants have a lot of characteristics that we’ve seen in seeds. They display protective desiccation tolerance mechanisms in their leaves, including anti-oxidants, protective proteins, and even dismantle their photosynthetic machinery during periods of drought. Even the cell wall structure and composition of resurrection plants resemble those of seeds. We are currently working on a paper describing the striking similarities between seeds and resurrection plants.
What was the most interesting discovery you made upon sequencing the genome of the resurrection plant?
First, the similarities between resurrection plants and seeds listed above were also apparent at the molecular level. For example, previous work suggested that the “ABI3 regulon”, consisting of about 100 genes regulated by the transcription factor ABI3, is specific to seeds, but we found that it is almost completely present (and active) in the leaves of Xerophyta viscosa too!
Secondly, we found “islands” or clusters of genes specific for desiccation tolerance that aren’t found in other species. Many of these regulate secondary metabolite pathways.
How challenging was it to sequence the genome of this plant? How did you overcome any difficulties?
It was very challenging. First, the species is an octoploid, meaning it has eight copies of each chromosome. This meant that we had to sequence its genome at very high coverage and employing the most advanced sequencing facilities, e.g. PacBio. Getting funding for this complex analysis was another challenge. We then took almost a year to assemble the genome and annotate it at the desired quality.
Xerophyta viscosa before and after the rains. Image credit: Prof. Henk Hilhorst.
You identified some of the most important genes involved in desiccation tolerance. Is it possible to translate this work into other species, such as crops that may be threatened by drought as the climate changes?
That will be our ultimate goal. It’s important to remember that desiccation-sensitive plants, including all our major crops, produce seeds that are desiccation tolerant. This implies that the information for desiccation tolerance is present in the genomes of these crops but that it is only turned on in the seeds. We are trying to determine how this is localized, in order to find a method to turn on the desiccation tolerance mechanism in vegetative parts of the (crop) plant too. In parallel we are expressing some of the key transcription factors from Xerophyta viscosa in some important crops to see how this affects them.
Are there any other interesting aspects of Xerophyta viscosa biology?
Contrary to plants that wilt and ultimately die because of (severe) drought, leaves of resurrection species do not show such stress-related senescence. This is related to the engagement of active anti-senescence genes during the drying of the leaves of resurrection species. We are currently investigating these senescence-related mechanisms too.
The rose of Jericho (Anastatica hierochuntica) is another resurrection plant. Image credit: FloraTrek. Used under license: CC BY-SA 3.0.
Do you expect to find that different types of desiccation-tolerant plants use the same subset of genes to survive drought, or could they have developed other pathways to resilience?
We expect that the core mechanism is very similar among the resurrection species but that each species may have adapted to its specific environment.
Funding permitting, we will sequence the genomes of at least another ten resurrection species to further clarify the various evolutionary pathways to desiccation tolerance and, importantly, to discriminate between species-specific and desiccation tolerance-specific genes.
What advice do you have for early career researchers?
Stick to what you believe in, even if you have to (temporarily) be involved in research that you appreciate less, e.g., because of better funding opportunities.
[MEXICO CITY] An international team of scientists identified a hundred genes that influence adaptation to the latitude, altitude, growing season and flowering time of nearly 4,500 native maize varieties in Mexico and in almost all Latin American and Caribbean countries.
Creole — or native — varieties of maize are derived from improvements made over thousands of years by local farmers, and contain genes that help them adapt to different environments.
“We are now using this analysis to find other genes that are of vital importance to breeders, such as those resistant to extreme heat, frost or drought — environmental conditions associated with climate change and that could affect maize production.”
Sarah Hearne, CIMMYT
“Latin American breeders will be able to use these results to identify native varieties that could contribute to improved adaptation”, Edward Buckler, a Cornell University researcher and co-author of the study published in Nature Genetics (February 6), told SciDev.Net.
The information on the genetic markers described in the study will be available online, said Sarah Hearne, a researcher at the International Maize and Wheat Improvement Center (CIMMYT) and co-author of the study. “Meanwhile, any breeder can contact us to request information”, she said.
“We are now using this analysis to find other genes that are of vital importance to breeders, such as those resistant to extreme heat, frost or drought — environmental conditions associated with climate change and that could affect maize production”, Hearne said.
Maize ears from CIMMYT’s collection, showing a wide variety of colors and shapes. CIMMYTs germplasm bank contains about 28,000 unique samples of cultivated maize and its wild relatives, teosinte and Tripsacum. These include about 26,000 samples of farmer landracestraditional, locally-adapted varieties that are rich in diversity. The bank both conserves this diversity and makes it available as a resource for breeding. Photo credit: Xochiquetzal Fonseca/CIMMYT.
Studying native maize varieties is extremely difficult because of their genetic variation. Although domesticated, they are wilder than commercial varieties.
For this study, the researchers cultivated hybrid creole varieties in various environments in Latin America and identified regions of the genome that control growth rates. They looked into where the varieties came from and what genetic features contributed to their growth in that environment.
In comments to SciDev.Net, James Holland, a researcher at North Carolina State University, Jeffrey Ross-Ibarra, a researcher at the University of California Davis, and Rodomiro Ortiz, a researcher at the Swedish University of Agricultural Sciences — who did not participate in the study — commended the magnitude of the study and the original method developed by the researchers to access the rich set of genetic information about native maize varieties.
Hearne added that the research team has initiated a “pre-breeding” programme with a small group of breeders in Mexico. As part of that programme, CIMMYT delivers to breeders materials from its germplasm bank of Creole maize; it also provides molecular information the breeders can use to generate new varieties.
This week’s post comes to us from Crystal Chan, project manager of the Application of Genomic Innovation in the Lentil Economy project led by Dr. Kirstin Bett at the Department of Plant Sciences, University of Saskatchewan.
Could you begin with a brief introduction to your research?
Our research focuses on the smart use of diverse genetic materials and wild relatives in the lentil (Lens culinaris) breeding program.
Canada has become the world’s largest producer and exporter of lentils in recent years. Lentils are an introduced species to the northern hemisphere and, until recently, our breeding program at the University of Saskatchewan involved just a handful of germplasms adapted to our climatic condition. With dedicated breeding efforts we have achieved noteworthy genetic gains in the past decade, but we are missing out on the vast genetic diversity available within the Lens genus. This is a major dilemma faced by all plant breeders: do we want consistency (sacrificing genetic diversity and reducing genetic gains over time) or diversity (sacrificing some important fixed traits and spending lots of time and resources in “backcrossing/rescue efforts”)?
In our current research, we use genomic tools to understand the genetic variability found in different lentil genotypes and the basis of what makes lentils grow well in different global environments (North America vs. Mediterranean countries vs. South Asian countries). We will then develop molecular breeding tools that breeders can use to improve the diversity and productivity of Canadian lentils while maintaining their adaptation to the northern temperate climate.
What first led you to this research topic?
Dr. Albert (Bert) Vandenberg, professor and lentil breeder at the University of Saskatchewan, noticed one of the wild lentil species was resistant to several diseases that devastate the cultivated lentil. After years of dedicated breeding effort, he was able to transfer the resistance traits to the cultivated lentil, but it took a lot of time and resources. We began looking into other beneficial traits and became fascinated with the domestication and adaptation aspects of lentil – after all lentil is one of the oldest cultivated crops, domesticated by man around 11,000 BC! With the rapid advance in genomic technology, we can start to better understand the biology and develop tools to harness these valuable genetic resources.
You have been involved in the development of tools that assist researchers to build databases of genomics and genetics data. Could you tell us more about projects such as Tripal?
Over the past six years, Lacey Sanderson (bioinformaticist in our group) has developed a database for our pulse research program at the University (Knowpulse, http://knowpulse.usask.ca/portal/). The database is specifically designed to present data that is relevant to breeders, as our group has a strong focus on variety development for the Canadian pulse crop industry. Knowpulse houses genotypic information from past and on-going lentil genomics projects, and includes tools for looking up genotypes as well as comparing the current genome assembly (currently v1.2) and other sequenced legume genomes. The tools are being developed in Tripal, an open-source toolkit that provides an interface between the data and a Drupal web content management system, in collaboration with colleagues at Washington State University.
At the moment we are developing new functionalities that will allow us to store and present germplasm information as well as phenotypic data. We are also working with our colleagues at Washington State University (under the “Tripal Gateway Project” funded by the National Science Foundation) to enhance interconnectivity between Knowpulse and other legume databases, such as the Legume Information Service (LIS) and Soybase, to facilitate comparative genomic studies.
How challenging are pulse genomes to assemble? How closely related are the various crops?
We had the fortune to lead the lentil genome sequencing initiative thanks to the support from producer groups and governments across the globe. The lentil genome is really challenging to assemble! We see nice synteny between lentil and the model legume, medicago, however the lentil genome is much bigger. We see a significant increase in genome size between chickpea and beans versus lentil (and pea for that matter), yet we have evidence to show that genome duplication is not the cause of the size increase. There are a lot of very long repetitive elements sprinkled around the genome, which makes its sequencing and proper assembly very challenging. Not to mention understanding the role of these long repetitive elements in biological functions…
What insights into crop domestication have you gained from these genomes?
That’s what we are working on right now under the AGILE (“Application of Genomics to Innovation in the Lentil Economy”) project. Stay tuned!
Do you work with breeders to develop new cultivars? What sorts of traits are most important?
Breeding is at the core of our work – both Kirstin and Bert are breeders (Kirstin has an active dry bean breeding program when she’s not busy with genomic research). All our research aims to feed information to the breeders so that they can make better crossing and selection decisions. Our work in herbicide tolerance has led to the development and implementation of a molecular marker to screen for herbicide resistance. With that marker we save time (skipping a crossing cycle) and forego the herbicide spraying test for all of our early materials.
Disease resistance and drought tolerance are also important for the growers. Visual quality (seed shape, size, color) are very important too as our customers are very picky as to what sort of lentils they like to buy/eat.
What does the future of legume/lentil agriculture hold?
Lentils have been a staple food in many countries for centuries and have been gaining popularity in North America in recent years as people are looking for plant-based protein sources. Lentils are high in fibre, protein, and complex carbohydrates, while low in fat and calories, and have a low glycemic index. They are suitable for vegetarian/vegan, gluten-free, diabetic, and heart-smart diets. Lentils also provide essential micronutrients such as iron, zinc and folates. Lentils are widely recognized as nutrient-dense food that could serve as part of the solution to combat global food and nutritional insecurity.
In modern agriculture, adding lentil or other leguminous crops in the crop rotation helps improve soil structure, soil quality, and biotic diversity, as well as enhancing soil fertility through their ability to fix nitrogen. Because pulse crops require little to no nitrogen fertilizer, they use half of the non-renewable energy inputs of other crops, reducing greenhouse gas emissions.
Sales of quinoa (Chenopodium quinoa) have exploded in the last decade, with prices more than tripling between 2008 and 2014. The popularity of this pseudocereal comes from its highly nutritious seeds, which resemble grains and contain a good balance of protein, vitamins, and minerals. The nourishing nature of quinoa meant it was prized by the Incas, who called it the “Mother grain”.
Quinoa is a popular ‘grain’, but it is more closely related to spinach and beetroot than cereals like wheat or barley. Image credit: Flickr user. Used under license: CC BY 2.0.
Quinoa is native to the Andes of South America, where it thrives in a range of conditions from coastal regions to alpine regions of up to 4000 m above sea level. Its resilience and nutritious seeds means that quinoa has been identified as a key crop for enhancing food security, but there are currently very few breeding programs targeting this species.
The challenge of improving the efficiency and sustainability of quinoa production has so far been restricted by the lack of a reference genome. This week, a team of researchers led by Professor Mark Tester (King Abdullah University of Science & Technology; KAUST) addressed this issue, publishing a high-quality genome sequence for quinoa in Nature. They compared the genome with that of related species to characterize the evolution and domestication of the crop, and investigated the genetic diversity of economically important traits.
The evolution of quinoa
Tester and colleagues used an array of genomics techniques to assemble 1.39 Gb of the estimated 1.45-1.50 Gb full length of quinoa’s genome. Quinoa is a tetraploid, meaning it has four copies of each chromosome. The researchers shed light on the evolutionary history of this crop by sequencing descendants of the two diploid species (each containing two sets of chromosomes) that hybridized to generate quinoa; kañiwa (Chenopodium pallidicaule) and Swedish goosefoot (Chenopodium suecicum). Comparing these sequences to quinoa and other relatives, the team showed that the hybridization event likely occurred between 3.3 and 6.3 million years ago. A comparison with other closely related Chenopodium species also suggested that, contrary to previous predictions, quinoa may have been domesticated twice, both in highland and coastal environments.
Quinoa seeds are coated with soap-like chemicals called saponins, which have a bitter taste that deters herbivores. Saponins can disrupt the cell membranes of red blood cells, so they have to be removed before human consumption, but this process is costly, so quinoa breeders are always looking for varieties that produce lower levels of saponins.
Sweet (low-saponin) quinoa strains do occur naturally, but the genes that regulate this phenotype were previously unknown. Tester and colleagues investigated sweet and bitter quinoa strains and discovered that a single gene (TRITERPENE SAPONIN BIOSYNTHESIS ACTIVATING REGULATOR-LIKE 1 [TSARL1]) controls the amount of saponins produced in the seeds. The low-saponin quinoa strains contained mutations in TSARL1 that prevented it from functioning properly. This is a key target for the improvement of quinoa in the future, although farmers will have to find new ways to protect their crops from birds and other seed predators!
The high-quality reference genome for quinoa generated by Tester and colleagues is likely to be vital for allowing many exciting improvements in the future. Breeders hoping to improve the yield, ease of harvest, stress tolerance, and saponin content of quinoa can develop genetic markers to speed up breeding for these key traits, improving the productivity of quinoa varieties and enhancing future food security.
Read the paper in Nature: Jarvis et al., 2017. The genome of Chenopodium quinoa. Nature. DOI: 10.1038/nature21370
Thank you to Professor Mark Tester (KAUST) for providing information used in this post!
Another fantastic year of discovery is over – read on for our 2016 plant science top picks!
A Zostera marina meadow in the Archipelago Sea, southwest Finland. Image credit: Christoffer Boström (Olsen et al., 2016. Nature).
The year began with the publication of the fascinating eelgrass (Zostera marina) genome by an international team of researchers. This marine monocot descended from land-dwelling ancestors, but went through a dramatic adaptation to life in the ocean, in what the lead author Professor Jeanine Olsen described as, “arguably the most extreme adaptation a terrestrial… species can undergo”.
One of the most interesting revelations was that eelgrass cannot make stomatal pores because it has completely lost the genes responsible for regulating their development. It also ditched genes involved in perceiving UV light, which does not penetrate well through its deep water habitat.
Plants are known to form new organs throughout their lifecycle, but it was not previously clear how they organized their cell development to form the right shapes. In February, researchers in Germany used an exciting new type of high-resolution fluorescence microscope to observe every individual cell in a developing lateral root, following the complex arrangement of their cell division over time.
Using this new four-dimensional cell lineage map of lateral root development in combination with computer modelling, the team revealed that, while the contribution of each cell is not pre-determined, the cells self-organize to regulate the overall development of the root in a predictable manner.
Watch the mesmerizing cell division in lateral root development in the video below, which accompanied the paper:
In March, a Spanish team of researchers revealed how the anti-wilting molecular machinery involved in preserving cell turgor assembles in response to drought. They found that a family of small proteins, the CARs, act in clusters to guide proteins to the cell membrane, in what author Dr. Pedro Luis Rodriguez described as “a kind of landing strip, acting as molecular antennas that call out to other proteins as and when necessary to orchestrate the required cellular response”.
In April, we received an amazing insight into the ‘decision-making ability’ of plants when a Swiss team discovered that plants can punish mutualist fungi that try to cheat them. In a clever experiment, the researchers provided a plant with two mutualistic partners; a ‘generous’ fungus that provides the plant with a lot of phosphates in return for carbohydrates, and a ‘meaner’ fungus that attempts to reduce the amount of phosphate it ‘pays’. They revealed that the plants can starve the meaner fungus, providing fewer carbohydrates until it pays its phosphate bill.
Author Professor Andres Wiemskenexplains: “The plant exploits the competitive situation of the two fungi in a targeted manner, triggering what is essentially a market-based process determined by cost and performance”.
The transition of ancient plants from water onto land was one of the most important events in our planet’s evolution, but required a massive change in plant biology. Suddenly plants risked drying out, so had to develop new ways to survive drought.
In May, an international team discovered a key gene in moss (Physcomitrella patens) that allows it to tolerate dehydration. This gene, ANR, was an ancient adaptation of an algal gene that allowed the early plants to respond to the drought-signaling hormone ABA. Its evolution is still a mystery, though, as author Dr. Sean Stevensonexplains: “What’s interesting is that aquatic algae can’t respond to ABA: the next challenge is to discover how this hormone signaling process arose.”
Sometimes revisiting old ideas can pay off, as a US team revealed in June. In 1930, Ernst Münch hypothesized that transport through the phloem sieve tubes in the plant vascular tissue is driven by pressure gradients, but no-one really knew how this would account for the massive pressure required to move nutrients through something as large as a tree.
Professor Michael Knoblauch and colleagues spent decades devising new methods to investigate pressures and flow within phloem without disrupting the system. He eventually developed a suite of techniques, including a picogauge with the help of his son, Jan, to measure tiny pressure differences in the plants. They found that plants can alter the shape of their phloem vessels to change the pressure within them, allowing them to transport sugars over varying distances, which provided strong support for Münch flow.
BLOG: We featured similar work (including an amazing video of the wound response in sieve tubes) by Knoblauch’s collaborator, Dr. Winfried Peters, on the blog – read it here!
Preserved remains of rope, seeds, reeds and pellets (left), and a desiccated barley grain (right) found at Yoram Cave in the Judean Desert. Credit: Uri Davidovich and Ehud Weiss.
In July, an international and highly multidisciplinary team published the genome of 6,000-year-old barley grains excavated from a cave in Israel, the oldest plant genome reconstructed to date. The grains were visually and genetically very similar to modern barley, showing that this crop was domesticated very early on in our agricultural history. With more analysis ongoing, author Dr. Verena Schünemannpredicts that “DNA-analysis of archaeological remains of prehistoric plants will provide us with novel insights into the origin, domestication and spread of crop plants”.
BLOG: We interviewed Dr. Nils Stein about this fascinating work on the blog – click here to read more!
Another exciting cereal paper was published in August, when an Australian team revealed that C4 photosynthesis occurs in wheat seeds. Like many important crops, wheat leaves perform C3 photosynthesis, which is a less efficient process, so many researchers are attempting to engineer the complex C4 photosynthesis pathway into C3 crops.
This discovery was completely unexpected, as throughout its evolution wheat has been a C3 plant. Author Professor Robert Henrysuggested: “One theory is that as [atmospheric] carbon dioxide began to decline, [wheat’s] seeds evolved a C4 pathway to capture more sunlight to convert to energy.”
Professor Stefan Jansson cooks up “Tagliatelle with CRISPRy fried vegetables”. Image credit: Stefan Jansson.
September marked an historic event. Professor Stefan Jansson cooked up the world’s first CRISPR meal, tagliatelle with CRISPRy fried vegetables (genome-edited cabbage). Jansson has paved the way for CRISPR in Europe; while the EU is yet to make a decision about how CRISPR-edited plants will be regulated, Jansson successfully convinced the Swedish Board of Agriculture to rule that plants edited in a manner that could have been achieved by traditional breeding (i.e. the deletion or minor mutation of a gene, but not the insertion of a gene from another species) cannot be treated as a GMO.
Phytochromes help plants detect day length by sensing differences in red and far-red light, but a UK-Germany research collaboration revealed that these receptors switch roles at night to become thermometers, helping plants to respond to seasonal changes in temperature.
Dr Philip Wiggeexplains: “Just as mercury rises in a thermometer, the rate at which phytochromes revert to their inactive state during the night is a direct measure of temperature. The lower the temperature, the slower phytochromes revert to inactivity, so the molecules spend more time in their active, growth-suppressing state. This is why plants are slower to grow in winter”.
A fossil ginkgo (Ginkgo biloba) leaf with its modern counterpart. Image credit: Gigascience.
In November, a Chinese team published the genome of Ginkgo biloba¸ the oldest extant tree species. Its large (10.6 Gb) genome has previously impeded our understanding of this living fossil, but researchers will now be able to investigate its ~42,000 genes to understand its interesting characteristics, such as resistance to stress and dioecious reproduction, and how it remained almost unchanged in the 270 million years it has existed.
Author Professor Yunpeng Zhaosaid, “Such a genome fills a major phylogenetic gap of land plants, and provides key genetic resources to address evolutionary questions [such as the] phylogenetic relationships of gymnosperm lineages, [and the] evolution of genome and genes in land plants”.
The year ended with another fascinating discovery from a Danish team, who used fluorescent tags and microscopy to confirm the existence of metabolons, clusters of metabolic enzymes that have never been detected in cells before. These metabolons can assemble rapidly in response to a stimulus, working as a metabolic production line to efficiently produce the required compounds. Scientists have been looking for metabolons for 40 years, and this discovery could be crucial for improving our ability to harness the production power of plants.
Genome editing technologies comprise a diverse set of molecular tools that allow the targeted modification of a DNA sequence within a genome. Unlike “traditional” breeding, genome editing does not rely on random DNA recombination; instead it allows the precise targeting of specific DNA sequences of interest. Genome editing approaches induce a double strand break (DSB) of the DNA molecule at specific sites, activating the cell’s DNA repair system. This process could be either error-prone, thus used by scientists to deactivate “undesired” genes, or error-free, enabling target DNA sequences to be “re-written” or the insertion of DNA fragments in a specific genomic position.
The most promising among the genome editing technologies, CRISPR/Cas9, was chosen as Science’s 2015 Breakthrough of the Year. Cas9 is an enzyme able to target a specific position of a genome thanks to a small RNA molecule called guide RNA (gRNA). gRNAs are easy to design and can be delivered to cells along with the gene encoding Cas9, or as a pre-assembled Cas9-gRNA protein-RNA complex. Once inside the cell, Cas9 cuts the target DNA sequence homologous to the gRNAs, producing DSBs.
The guide RNA (sgRNA) directs Cas9 to a specific region of the genome, where it induces a double-strand break in the DNA. On the left, the break is repaired by non-homologous-end joining, which can result in insertion/deletion (indel) mutations. On the right, the homologous-directed recombination pathway creates precise changes using a supplied template DNA. Credit: Ran et al. (2013). Nature Protocols.
Genome editing in crops
Together with the increased data availability on crop genomes, genome editing techniques such as CRISPR are allowing scientists to carry out ambitious research on crop plants directly, building on the knowledge obtained during decades of investigation in model plants.
The concept of CRISPR was first tested in crops by generating cultivars that are resistant to herbicides, as this is an easy trait to screen for and identify. One of the first genome-edited crops, a herbicide-resistant oilseed rape produced by Cibus, has already been grown and harvested in the USA in 2015.
Researchers used CRISPR to engineer a wheat variety resistant to powdery mildew (shown here), a major disease of this crop. Image credit: NY State IPM Program. Used under license: CC BY 2.0.
Genome editing could also revolutionize the management of viral plant disease. The CRISPR/Cas9 system was originally discovered in bacteria, where it provided them with molecular immunity against viruses, but it can also be moved into plants. Scientists can transform plants to produce the Cas9 and gRNAs that target viral DNA, reducing virus accumulation; alternatively, they can suppress those plant genes that are hijacked by the virus to mediate its own diffusion in the plants. Since most plants are defenseless against viruses and there are no chemical controls available for plant viruses, the main method to stop the spread of these diseases is still the destruction of the infected plant. For the first time in history, scientists have an effective weapon to fight back against plant viruses.
The cassava brown streak disease virus can destroy cassava crops, threatening the food security of the 300 million people who rely on this crop in Africa. Image credit: Katie Tomlinson (for more on this topic, read her blog here).
Genome editing will be particularly useful in the genetic improvement of many crops that are propagated mainly by vegetative reproduction, and so very difficult to improve by traditional breeding methods involving crossing (e.g. cassava, banana, grape, potato). For example, using TALENs, scientists from Cellectisedited a potato line to minimize the accumulation of reducing sugars that may be converted into acrylamide (a possible carcinogen) during cooking.
Concerns about off-targets
One of the hypothesized risks of using CRISPR/Cas9 is the potential targeting of undesired DNA regions, called off-targets. It is possible to limit the potential for off-targets by designing very specific gRNAs, and all of the work published so far either did not detect any off-targets or, if detected, they occurred at a very low frequency. The number of off-target mutations produced by CRISPR/Cas9 is therefore minimal, especially if compared with the widely accepted random mutagenesis of crops used in plant breeding since the 1950s.
GM or not-GM
Genome editing is interesting from a regulatory point of view too. After obtaining the desired heritable mutation using CRISPR/Cas9, it is possible to remove the CRISPR/Cas9 integrated vectors from the genome using simple genetic segregation, leaving no trace of the genome modification other than the mutation itself. This means that some countries (including the USA, Canada, and Argentina) consider the products of genome editing on a case-by-case basis, ruling that a crop is non-GM when it contains gene combinations that could have been obtained through crossing or random mutation. Many other countries are yet to issue an official statement on CRISPR, however.
Recently, scientists showed that is possible to edit the genome of plants without adding any foreign DNA and without the need for bacteria- or virus-mediated plant transformation. Instead, a pre-assembled Cas9-gRNA ribonucleoprotein (RNP) is delivered to plant cells in vitro, which can edit the desired region of the genome before being rapidly degraded by the plant endogenous proteases and nucleases. This non-GM approach can also reduce the potential of off-target editing, because of the minimal time that the RNP is present inside the cell before being degraded. RNP-based genome editing has been already applied to tobacco plants, rice, and lettuce, as well as very recently to maize.
In conclusion, genome editing techniques, and CRISPR/Cas9 in particular, offers scientists and plant breeders a flexible and relatively easy approach to accelerate breeding practices in a wide variety of crop species, providing another tool that we can use to improve food security in the future.
For more on CRISPR, check out this recent TED Talk from Ellen Jorgensen:
About the author
Dr Damiano Martignago is a plant molecular biologist who graduated from Padua University, Italy, with a degree in Food Biotechnology in 2009. He obtained his PhD in Biology at Roma Tre University in 2014. His experience with CRISPR/Cas9 began in the lab of Prof. Fabio Fornara (University of Milan), where he used CRISPR/Cas9 to target photoperiod genes of interest in rice and generate mutants that were not previously available. He recently moved to Rothamsted Research, UK, where he works as Genome Editing Specialist, transferring CRISPR/Cas9 technology to hexaploid bread wheat with the aim of improving the efficiency of genome editing in this crop. He is actively involved with AIRIcerca (International Association of Italian Scientists), disseminating and promoting scientific news.
The Atacama Desert is a strip of land near 1000 km in length located in northern Chile. With an average yearly rainfall of just 15 mm (close to 0 in some locations) and high radiation levels, it is the driest desert in the world. Geological estimates suggest that the Atacama has remained hyperarid for at least eight million years. Standing in its midst, one may easily feel as though visiting a valley on Mars.
Despite these harsh environmental conditions, it is possible to find life in the Atacama. At the increased altitudes along the western slopes of the Andes precipitation is slightly increased, allowing plant life.
The driest and oldest desert in the world acts as a natural laboratory where for 150 million years plants adapted to and colonized this environment. These adaptations are likely present in multiple desert plant lineages, thus providing an evolutionary framework where these traits can be associated with a signature of convergent evolution.
Surviving a nitrogen-limited landscape
Image credit: Center for Genome Regulation
The interplay of environmental conditions in the transect of the Atacama, ranging from 2500 to 4500 meters above sea level, results in three broad microclimates; Pre Puna, Puna, and High Steppe. These microclimates have different humidities, temperatures, levels of organic matter and even different pH levels, but share one common feature: low nitrogen levels.
To engineer crops with higher nitrogen use efficiency, it is very useful to first learn how plants adapt to growth in low nitrogen environments. Here the Atacama Desert enters into the game. Plants growing in the desert can survive 100-fold less nitrogen below optimum concentrations. Using phylogenetics it is possible to uncover novel genes and mechanisms related to adaptation to these extreme conditions, which have not been discovered through traditional genetic approaches.
Currently, nitrogen fertilizers are widely employed to increase crop yield. In 2008 100 million tons of this fertilizer were used and it is projected that for 2018 the demand for nitrogen will rise to 119 million tons. Regretfully, the production and over-usage of this type of fertilizer has an enormous impact in the environment and human health. Around 60% of the nitrogen introduced to the soil for agricultural purposes is leached and lost. Moreover, nitrogen runoffs to the water cause eutrophication in both freshwater and marine ecosystems, leading to algae and phytoplankton blooms, low levels of dissolved oxygen, and finally the migration or death of the present fauna, forming dead zones such as the one in the Gulf of Mexico.
Image credit: Center for Genome Regulation
Nitrogen fertilizers are not the only major concern in modern agricultural procedures. The co-localization of drought and low nitrogen levels is especially detrimental for plant growth and development. We need to support not only the nutritional requirement of an expanding global population but also new energetic strategies based on production of biomass for biofuels on marginal nutrient poor soils. In order to increase crop yields while reducing the environmental impact of nitrogen fertilizers, it is necessary to develop new agricultural strategies and cutting edge technologies.
Learning from the desert
What if we could profit from the extraordinary plants that have had thousands of years to learn how to cope with nitrogen scarcity, drought and extreme radiation? Specifically, can we unravel the genes and mechanisms that allow them to survive in such a barren place?
Image credit: Center for Genome Regulation
Over the past three years our group has identified 62 different plant species that inhabit the Atacama Desert, and established a correlation between their habitat attributes and biological characteristics. Using tools such as whole transcriptome shotgun sequencing or RNA-Seq complemented with different bioinformatics approaches, we have identified over 896,000 proteins that are expressed in these conditions.
In this way we aim to learn which processes are highly utilized in these “extreme survivors” compared to similar species that are present in the deserts of California, where the climatic conditions are similar but there is no nitrogen scarcity. That is how we expect to find new mechanisms (or, more precisely, very old mechanisms) that enable plants to survive and grow efficiently in extreme environments.
With another year nearly over we recently put out a call for nominations for the Most Influential Plant Science Research of 2015. Suggestions flooded in, and we also trawled through our social media feeds to see which stories inspired the most discussion and engagement. It was fantastic to read about so much amazing research from around the world. Below are our top five, selected based on impact for the plant science research community, engagement on social media, and importance for both policy and potential end product/application.
Choosing the most inspiring stories was not an easy job. If you think we’ve missed something, please let us know in the comments below, or via Twitter! In the coming weeks we’ll be posting a 2015 Plant Science Round Up, which will include other exciting research that didn’t quite make the top five, so watch this space!
Scientists at the International Potato Center in Lima, Peru, found that 291 varieties of sweet potato actually contain bacterial genes. This technically means that sweet potato is a naturally occurring genetically modified crop! Alongside all the general discussion about GM regulations, particularly in parts of Europe where regulations about growing GM crops have been decentralized from Brussels to individual EU Member States, this story caused much discussion on social media when it was published in March of this year.
It is thought that ancestors of the modern sweet potato were genetically modified by bacteria in the soil some 8000 years ago. Scientists hypothesize that it was this modification that made consumption and domestication of the crop possible. Unlike the potato, sweet potato is not a tuber but a mere root. The bacteria genes are thought to be responsible for root swelling, giving it the fleshy appearance we recognize today.
RNA-guided Cas9 nuclease creates targetable heritable mutations in Barley and Brassica
Our number two on the list also relates to genetic modification, this time focusing on methods. Regardless of whether or not we want to have genetically modified crops in our food supply, GM is a valuable tool used by researchers to advance knowledge of gene function at the genetic and phenotypic level. Therefore, systems of modification that make the process faster, cheaper, and more accurate provide fantastic opportunities for the plant science community to progress its understanding.
The Cas9 system is a method of genome editing that can make precise changes at specific locations in the genome relatively cheaply. This novel system uses small non-coding RNA to direct Cas9 nuclease to the DNA target site. This type of RNA is small and easy to program, providing a flexible and easily accessible system for genome editing.
Inheritance of genome modifications using Cas9 has previously been shown in the model plants, Arabidopsis and rice. However, the efficiency of this inheritance, and therefore potential application in crop plants has been questionable.
The breakthrough study published in November by researchers at The Sainsbury Laboratory and John Innes Centre both in Norwich, UK, demonstrated the mutation of two commercial crop plants, Barley and Brassica oleracea, using the Cas9 system and subsequent inheritance mutations.
This is an incredibly exciting development in the plant sciences and opens up many options in the future in terms of genome editing and plant science research.
Striga is a parasitic plant that mainly affects parts of Africa. It is a major threat to food crops such as rice and corn, leading to yield losses worth over 10 billion US dollars, and affecting over 100 million people.
Striga infects the host crop plant through its roots, depriving them of their nutrients and water. The plant hormone strigolactone, which is released by host plants, is known to induce Striga germination when host plants are nearby.
In a study published in August of this year the Striga receptors for this hormone, and the proteins responsible for striga germination were identified.
Striga plants are known to wither and die if they cannot find a host plant upon germination. Induction of early germination using synthetic hormones could therefore remove Striga populations before crops are planted. This work is vital in terms of regulating Striga populations in areas where they are hugely damaging to crop plants and people’s livelihoods.
Resurrection plants are a unique group of flora that can survive extreme water shortages for months or even years. There are more than 130 varieties in the world, and many researchers believe that unlocking the genetic codes of drought-tolerant plants could help farmers working in increasingly hot and dry conditions.
During a drought, the plant acts like a seed, becoming so dry that it appears dead. But as soon as the rains come, the shriveled plant bursts ‘back to life’, turning green and robust in just a few hours.
O. thomaeum is a small C4 grass species found in Africa and India. It is closely related to major food feed and bioenergy crops. Therefore this work represents a significant step in terms of understanding novel drought tolerance mechanisms that could be used in agriculture.
Supercomputing overcomes major ecological challenge
Currently, one of the greatest challenges for ecologists is to quantify plant diversity and understand how this affects plant survival. For the last 500 years independent research groups around the world have collected this diversity data, which has made organization and collaboration difficult in the past.
There are approximately 120,000 plant species in North and South America, but mapping and determining the hotspots of species richness requires computationally intensive geographic range estimates. With supercomputing the BIEN group could generate and store geographic range estimates for plant species in the Americas.
It also gives ecologists the ability to document continental scale patterns of species diversity, which show where any species of plant might be found. These novel maps could prove a fantastic resource for ecologists working on diversity and conservation.
Read more about this story on the TACC website, here.
Do you have a genome sequencer in your pocket or are you just happy to see me?
By Nikolai Adamski
On September 4 I attended an event sponsored by Oxford Nanopore Technologies (ONT) at Norwich Research Park, UK, which focused on nanopore technologies. This new technology has been dubbed ‘Next, next-generation sequencing’, and could have really exciting implications for the future of genome sequencing.
ONT has developed a pocked-sized genome sequencing device called the MinION that can sequence genomes in real time. Thanks to recent pop culture this generates visions of cuddly yellow creatures with an overly developed desire to serve super-villains. However, a MinION is actually a new genome sequencing device. To help confused readers, the figure below should help clarify the issue once and for all (Figure 1).
Figure 1: Demonstrating the difference between the pop culture Minion on the left and the genome sequencing MinION on the right.
The striking thing about the MinION is its size. Sequencing machines these days vary in size from something that sits on a desktop, to something that fills half a student’s room. The MinION however, fits in the palm of your hand. This is possible thanks to highly miniaturized electronics.
So how does it work?
At the core of the MinION are two biological components: the nanopore protein, which gives the company its name, and a motor protein. The nanopore protein sits on top of an artificial layer and acts a microscopic sluice gate that controls how much of the sample solution passes through it into the lower layer. The sample solution contains DNA, but also ions that pass through the nanopore, thus creating a measurable electrical current. If a big molecule like a strand of DNA passes through the nanopore, the flow of ions is perturbed, which results in a change in the electrical current. These changes are recorded and interpreted to give the sequence of said DNA molecule.
Meanwhile, the motor protein sticks to a DNA molecule, attaches itself to the top of the nanopore, and feeds the DNA through the nanopore as a single strand at a certain speed. This process is similar to a ratchet. Each MinION device has thousands of nanopores allowing for as many molecules to pass through and be sequenced in real time. This is nicely illustrated in a video made by ONT, which you can see here which is well worth a watch!
The sequence data are sent to a cloud server in real time, where they are transformed and analyzed and the final data sent back to the user. This eliminates the need for an expensive computer infrastructure as well as the need for extensive training in bioinformatics.
Limitations of the technology
So far so good, but there are still some issues with the MinION system. One of these is the average length of the DNA molecules that can be sequenced. In theory, the MinION system is able to sequence DNA molecules of any length, although the data from users at last week’s event suggests that, at the moment, the average length of sequence obtained is around 6,000 base pairs (bp). This is still a great value, but there is room for improvement. Another issue is the amount of data generated by a single MinION run, which according to user experience is generally around 1Gb, approximately 200 times the size of the gut bacterium E. coli. Both of these issues can be easily remedied by running several MinION sequencers with the same sample.
A larger problem is the matter of sequencing accuracy, which is now somewhere around 90%, although it can be as low as 75%. This can in part be compensated for by the sheer amount of data generated. However, it would require a lot of sequencing to make up for these mistakes, and is a critical point that needs to be addressed by ONT in the future.
There are many more fascinating applications for the MinION sequencer. Scientists who are interested can join the MinION Access Programme (MAP) to become part of the research and development community.
I very much enjoyed the ONT event and I am hopeful and curious about what the next few years will bring in terms of innovation and development.
Nikolai Adamski is a postdoctoral scientist working at the John Innes Centre in Norwich, UK, in the group of Cristobal Uauy. He studies yield and yield-related traits in wheat, trying to identify the underlying genes to understand the control and regulation of these traits.