“Once you’ve cloned a major gene in one crop, it is possible to find a counterpart gene that has the same function in another crop, and this is easier than finding useful genes from scratch” explains Leon Kochian, Professor in Plant Biology and Crop and Soil Sciences at Cornell University, USA, and Director of the Robert W Holley Center for Agriculture and Health, United States Department of Agriculture – Agricultural Research Service.
Aluminium toxicity and low phosphorus levels in acid soils are major factors that hinder cereal productivity worldwide, particularly in sub-Saharan Africa, South America and Southeast Asia. Globally, acid soils are outranked only by drought when it comes to stresses that threaten food security.
Tolerance to high levels of aluminium and low phosphorus is conferred by major genes, which lend themselves to cloning. Major genes are genes that by themselves have a significant and evident effect in producing a particular trait; it’s therefore easier to find and deploy a major gene associated with a desired trait than having to find and clone several minor genes.
Harvesting sorghum in Kenya.
Cloning major genes instrumental in hunt for resilient varieties
Locating a single gene within a plant’s DNA is like looking for a needle in a haystack. Instead of searching for a gene at random, geneticists need a plan for finding it.
The first step is to conduct prolonged phenotyping – that is, measuring and recording of plants’ observable characteristics in the field. By coupling and comparing this knowledge with genetic sequencing data, scientists can identify and locate quantitative trait loci (QTLs) – discrete genetic regions that contain genes associated with a desired trait, in this instance tolerance to aluminium or improved phosphorus uptake. They then dissect the QTL to single out the gene responsible for the desired trait. In the case of sorghum, researchers had identified the aluminium tolerance locus AltSB, and were looking for the gene responsible.
Once the gene has been identified, the next step is to clone it – that is, make copies of the stretch of DNA that makes up the gene. Geneticists need millions of copies of the same gene for their research: to gain information about the nucleotide sequence of the gene, create molecular markers to help identify the presence of the gene in plants and help compare versions of the gene from different species, and understand the mechanisms it controls and ways it interacts with other genes.
Drying the sorghum harvest in India.
Sorghum was one of the simpler crops to work with, according to Claudia Guimarães.
“Sorghum has a smaller genome… with clear observable traits, which are often controlled by one major gene,” she says.
The first breakthrough was the identification and cloning of SbMATE, the aluminium tolerance gene in sorghum behind the AltSB locus. The next was finding a diagnostic marker for the gene so that it could be used in breeding.
Marking genes to quickly scan plants for desired traits
Harvesting rice in The Philippines.
Once a desired gene is identified, a specific molecular marker must be found for it. This is a variation in the plant’s DNA, associated with a gene of interest, that scientists can use to flag up the gene’s presence. We can compare this process to using a text highlighter in a book, where the words represent the genes making up a genome. Each marker is like a coloured highlighter, marking sentences (genomic regions) containing particular keywords (genes) and making them easier to find.
In molecular breeding, scientists can use markers to quickly scan hundreds or thousands of DNA samples of breeding materials for a gene, or genes, that they want to incorporate into new plant varieties. This enables them to select parents for crosses more efficiently, and easily see which of the next generation have inherited the gene. This marker-assisted breeding method can save significant time and money in getting new varieties out into farmers’ fields.
Leon, who was also the Principal Investigator for various GCP-funded research projects, says that the cloning of SbMATE helped advance sorghum as a model for the further exploration of aluminium tolerance and the discovery of new molecular solutions for improving crop yields.
“This research also has environmental implications for badly needed increases in food production on marginal soils in developing countries,” says Leon. “For example, if we can increase food production on existing lands, it could limit agriculture’s encroachment into other areas.”
Rice field trials in Tanzania.
Aluminium toxicity is the result of aluminum becoming more available to plants when the soil pH is lower, and affects 38 percent of farmland in Southeast Asia, 31 percent in South America and 20 percent in East Asia, sub-Saharan Africa and North America. Meanwhile phosphorus, the second most important inorganic plant nutrient after nitrogen, becomes less available to plants in acid soils because it binds with aluminium and iron oxides. Almost half of the ricelands across the globe are currently phosphorus deficient. The research therefore has the potential for significant impact across the world.
The GCP-funded scientists used markers to search rice and maize for genes equivalent to sorghum’s SbMATE. In maize they successfully identified a similar gene, ZmMATE1, which is now being validated in Brazil, Kenya and Mali. In rice the search continues, but will become easier now that markers for ZmMATE1 have been developed.
Similarly, having validated, cloned and developed markers for PSTOL1 gene in rice, researchers at IRRI and JIRCAS then worked with researchers at EMBRAPA and Cornell University to use PSTOL1 markers to search for comparable genes in sorghum and maize. In both crops, genes similar to PSTOL1 have been identified and shown to improve grain yields under low-phosphorus soil conditions, albeit through different mechanisms.
The GCP-funded discoveries are already being used in marker-assisted selection in national breeding programmes in Brazil, Kenya, Niger, Indonesia, Japan, The Philippines and USA in sorghum, maize and rice. They have led to the release of new, aluminium-tolerant sorghum varieties in Brazil, with more currently being developed, along with phosphorus-efficient rice varieties.
Showing off freshly harvested sorghum in Kenya.
Cloning a worthwhile investment
Gene cloning was a relatively small cost in GCP’s research budget – about five percent (approximately USD 7 million) of a total research budget of USD 150 million spread over 10 years.
The gene-cloning component nonetheless yielded important genes for aluminium tolerance and phosphorus-uptake efficiency, within and across genomes. The molecular markers that have been developed are helping plant breeders across the world produce improved crop varieties.
Jean-Marcel Ribaut, Director of GCP, concludes: “The new markers developed for major genes in rice, sorghum and maize will have a significant impact on plant-breeding efficiency in developing countries.
“Breeders will be able to identify aluminium-tolerance and phosphorus-efficiency traits quicker, which, in time, will enable them to develop new varieties that will survive and thrive in the acid soils that make up more than half of the world’s arable soils.”
Maize is a staple crop for Kenyans, with 90 percent of the population depending on it for food. However, acid soils cause yield losses of 17–50 percent across the nation.
Soil acidity is a major environmental and economic concern in many more countries around the world. The availability of nutrients in soil is affected by pH, so acid conditions make it harder for plants to get a balanced diet. High acidity causes two major problems: perilously low levels of phosphorus and toxically high levels of aluminium. Aluminium toxicity affects 38 percent of farmland in Southeast Asia, 31 percent in Latin America and 20 percent in East Asia, sub-Saharan Africa and North America.
Aluminium toxicity in soil comes close to rivalling drought as a food-security threat in critical tropical food-producing regions. By damaging roots, acid soils deprive plants of the nutrients and water they need to grow – a particularly bitter effect when water is scarce.
Maize, meanwhile, is one of the most economically important food crops worldwide. It is grown in virtually every country in the world, and it is a staple food for more than 1.2 billion people in developing countries across sub-Saharan Africa and Latin America. In many cultures it is consumed primarily as porridge: polenta in Italy; angu in Brazil; and isitshwala, nshima, pap, posho,sadza or ugali in Africa.
Ugali, a stiff maize porridge that is a staple dish across East Africa, being prepared in Tanzania.
Maize is also a staple food for animals reared for meat, eggs and dairy products. Around 60 percent of global maize production is used for animal feed.
The world demand for maize is increasing at the same time as global populations burgeon and climate changes. Therefore, improving the ability of maize to withstand acid soils and produce higher yields with less reliable rainfall is paramount. This is why the CGIAR Generation Challenge Programme (GCP) invested almost USD 12.5 million into maize research between 2004 and 2014.
GCP’s goal was to facilitate the use of genetic diversity and advanced plant science to improve food security in developing countries through the breeding of ‘super’ crops – including maize – able to tolerate drought and poor soils and resist diseases.
Researchers take on the double whammy of acid soils and drought
Part of successfully breeding higher-yielding drought-tolerant maize varieties involves improving plant genetics for acid soils. In these soils, aluminium toxicity inhibits root growth, reducing the amount of water and nutrients that the plant can absorb and compounding the effects of drought.
Improving plant root development for aluminium tolerance and phosphorous efficiency can therefore have the positive side effect of higher plant yield when water is limited.
A farmer in Tanzania shows the effects of drought on her maize crop. The maize ears are undersized with few grains.
Although plant breeders have exploited the considerable variation in aluminium tolerance between different maize varieties for many years, aluminium toxicity has been a significant but poorly understood component of plant genetics. It is a particularly complex trait in maize that involves multiple genes and physiological mechanisms.
The solution is to take stock of what maize germplasm is available worldwide, characterise it, clone the sought-after genes and implement new breeding methods to increase diversity and genetic stocks.
Marilyn Warburton, then a molecular geneticist at CIMMYT, led this GCP-funded project. Her goal was to discover how all the genetic diversity in maize gene-bank collections around the globe might be used for practical plant improvement. She first gathered samples from gene banks all over the world, including those of CIMMYT and the International Institute of Tropical Agriculture (IITA). Scientists from developing country research centres in China, India, Indonesia, Kenya, Nigeria, Thailand and Vietnam also contributed by supplying DNA from their local varieties.
Researchers then used molecular markers and a bulk fingerprinting method – which Marilyn was instrumental in developing – for three purposes: to characterise the structure of maize populations, to better understand how maize migrated across the world, and to complete the global picture of maize biodiversity. Scientists were also using markers to search for new genes associated with desirable traits.
Allen Oppong, a maize pathologist and breeder from Ghana’s Crops Research Institute (CRI), of the Council for Scientific and Industrial Research, was supported by GCP from 2007 to 2010 to characterise Ghana’s maize germplasm. Trained in using the fingerprinting technique, Allen was able to identify distinctly different maize germplasm in the north of Ghana (with its dry savanna landscape) and in the south (with its high rainfall). He also identified mixed germplasm, which he says demonstrates that plant germplasm often finds its way to places where it is not suitable for optimal yield and productivity. Maize yields across the country are low.
Stocktaking a world’s worth of maize for GCP was a challenge, but not the only one, according to Marilyn. “In the first year it was hard to see how all the different partners would work together. Data analysis and storage was the hardest; everyone seemed to have their own idea about how the data could be stored, accessed and analysed best.
“The science was also evolving, even as we were working, so you could choose one way to sequence or genotype your data, and before you were even done with the project, a better way would be available,” she recalls.
Maize ears drying in Ghana.
Comparing genes: sorghum gene paves way for maize aluminium tolerance
In parallel to Marilyn’s work, scientists at the Brazilian Corporation of Agricultural Research (EMBRAPA) had already been advancing research on plant genetics for acid soils and the effects of aluminium toxicity on sorghum – spurred on by the fact that almost 70 percent of Brazil’s arable land is made up of acid soils.
What was of particular interest to GCP in 2004 was that the Brazilians, together with researchers at Cornell University in the USA, had recently mapped and identified the major sorghum aluminium tolerance locus AltSB, and were working on isolating the major gene within it with a view to cloning it. Major genes were known to control aluminium tolerance in sorghum, wheat and barley and produce good yields in soils that had high levels of aluminium. The gene had also been found in rape and rye.
GCP embraced the opportunity to fund more of this work with a view to speeding up the development of maize – as well as sorghum and rice – germplasm that can withstand the double whammy of acid soils and drought.
Maize trials in the field at EMBRAPA. The maize plants on the left are aluminium-tolerant and so able to withstand acid soils, while those on the right are not.
Leon Kochian, Director of the Robert W Holley Center for Agriculture and Health, United States Department of Agriculture – Agricultural Research Service and Professor at Cornell University, was a Principal Investigator for various GCP research projects investigating how to improve grain yields of crops grown in acid soils. “GCP was interested in our work because we were working with such critical crops,” he says.
“The idea was to use discoveries made in the first half of the GCP’s 10-year programme – use comparative genomics to look into genes of rice and maize to see if we can see relations in those genes – and once you’ve cloned a gene, it is easier to find a gene that can work for other crops.”
By combing the maize genome searching for a similar gene to sorghum’s SbMATE, Jurandir’s EMBRAPA colleague Claudia Guimarães and a team of GCP-supported scientists discovered the maize aluminium tolerance gene ZmMATE1. High expression of this gene, first observed in maize lines with three copies of ZmMATE1, has been shown to increase aluminium tolerance. ZmMATE1 improves grain yields in acid soil by up to one tonne per hectare when introgressed in an aluminium-sensitive line.
The genetic region, or locus, containing the ZmMATE1 aluminium tolerance gene is known as qALT6. Photo 1 shows a rhyzobox containing two layers of soil: a corrected top-soil and lower soils with 15 percent aluminium saturation. On the right, near-isogenic lines (NILs) introgressed with qALT6 show deeper roots and longer secondary roots in the acidic lower soil, whereas on the left the maize line without qALT6, L53, shows roots mainly confined to the corrected top soil. Photo 2 shows maize ears from lines without qALT6 (above) and with qALT6 (below); the lines with qALT6 maintain their size and quality even under high aluminium levels of 40 percent aluminium saturation.
The outcomes of these GCP-supported research projects provided the basic materials, such as molecular markers and donor sources of the positive alleles, for molecular-breeding programmes focusing on improving maize production and stability on acid soils in Latin America, Africa and other developing regions.
Kenya deploys powerful maize genes
One of those researchers crucial to achieving impact in GCP’s work in maize was Samuel (Sam) Gudu of Moi University, Kenya. From 2010 he was the Principal Investigator for GCP’s project on using marker-assisted backcrossing (MABC) to improve aluminium tolerance and phosphorous efficiency in maize in Kenya. This project combined molecular and conventional breeding approaches to speed up the development of maize varieties adapted to the acid soils of Africa, and was closely connected to the other GCP comparative genomics projects in maize and sorghum.
MABC is a type of marker-assisted selection (see box), which Sam’s team – including Dickson Ligeyo of KALRO – used to combine new molecular materials developed through GCP with Kenyan varieties. They have thus been able to significantly advance the breeding of maize varieties suitable for soils in Kenya and other African countries.
Maize and Comparative Genomics were two of seven Research Initiatives (RIs) where GCP concentrated on advancing researchers’ and breeders’ skills and resources in developing countries. Through this work, scientists have been able to characterise maize germplasm using improved trait observation and characterisation methods (phenotyping), implement molecular-breeding programmes, enhance strategic data management and build local human and infrastructure capacity.
The ultimate goal of the international research collaboration on comparative genomics in maize was to improve maize yields grown on acidic soils under drought conditions in Kenya and other African countries, as well as in Latin America. Seven institutes partnered up to for the comparative genomics research: Moi University, KALRO, EMBRAPA, Cornell University, the United States Department of Agriculture (USDA), JIRCAS and the International Rice Research Institute (IRRI).
“Before funding by GCP, we were mainly working on maize to develop breeding products resistant to disease and with increased yield,” says Sam. “At that time we had not known that soil acidity was a major problem in the parts of Kenya where we grow maize and sorghum. GCP knew that soil acidity could limit yields, so in the work with GCP we managed to characterise most of our acid soils. We now know that it was one of the major problems for limiting the yield of maize and sorghum.
“The relationship to EMBRAPA and Cornell University is one of the most important links we have. We developed material much faster through our collaboration with our colleagues in the advanced labs. I can see that post-GCP we will still want to communicate and interact with our colleagues in Brazil and the USA to enable us to continue to identify molecular materials that we discover,” he says. Sam and other maize researchers across Kenya, including Dickson, have since developed inbred, hybrid and synthetic varieties with improved aluminium tolerance for acid soils, which are now available for African farmers.
A Kenyan maize farmer.
“We crossed them [the new genes identified to have aluminium tolerance] with our local material to produce the materials we required for our conditions,” says Sam.
“The potential for aluminium-tolerant and phosphorous-efficient material across Africa is great. I know that in Ethiopia, aluminium toxicity from acid soil is a problem. It is also a major problem in Tanzania. It is a major problem in South Africa and a major problem in Kenya. So our breeding work, which is starting now to produce genetic materials that can be used directly, or could be developed even further in these other countries, is laying the foundation for maize improvement in acid soils.”
Sam is very proud of the work: “Several times I have felt accomplishment, because we identified material for Kenya for the first time. No one else was working on phosphorous efficiency or aluminium tolerance, and we have come up with materials that have been tested and have become varieties. It made me feel that we’re contributing to food security in Kenya.”
Maize grain for sale.
Maize for meat: GCP’s advances in maize genetics help feed Asia’s new appetites
Reaping from the substantial advances in maize genetics and breeding, researchers in Asia were also able to enhance Asian maize genetic resources.
A pig roots among maize ears on a small farm in Nepal.
Bindiganavile Vivek, a senior maize breeder for CIMMYT based in India, has been working with GCP since 2008 on improving drought tolerance in maize, especially for Asia, for two reasons: unrelenting droughts and a staggering growth the importance of maize as a feedstock. This work was funded by GCP as part of its Maize Research Initiative.
“People’s diets across Asia changed after government policies changed in the 1990s. We had a more free market economy, and along with that came more money that people could spend. That prompted a shift towards a non vegetarian diet,” Vivek recounts.
“Maize, being the number one feed crop of the world, started to come into demand. From the year 2000 up to now, the growing area of maize across Asia has been increasing by about two percent every year. That’s a phenomenal increase. It’s been replacing other crops – sorghum and rice. There’s more and more demand.
“Seventy percent of the maize that is produced in Asia is used as feed. And 70 percent of that feed is poultry feed.”
In Vietnam, for example, the government is actively promoting the expansion of maize acreage, again displacing rice. Other Asian nations involved in the push for maize include China, Indonesia and The Philippines.
A farmer in Indonesia transports his maize harvest by motorcycle.
The problem with this growth is that 80 percent of the 19 million hectares of maize in South and Southeast Asia relies on rain as its only source of water, so is prone to drought: “Wherever you are, you cannot escape drought,” says Vivek. And resource-poor farmers have limited access to improved maize products or hybrids appropriate for their situation.
Vivek’s research for GCP focused on the development – using marker-assisted breeding methods, specifically marker-assisted recurrent selection (MARS) – of new drought-tolerant maize adapted to many countries in Asia. His goal was to transfer the highest expression of drought tolerance in maize into elite well-adapted Asian lines targeted at drought-prone or water-constrained environments.
Asia’s existing maize varieties had no history of breeding for drought tolerance, only for disease resistance. To make a plant drought tolerant, many genes have to be incorporated into a new variety. So Vivek asked: “How do you address the increasing demand for maize that meets the drought-tolerance issue?”
The recent work on advancing maize genetics for acid soils in the African and Brazilian GCP projects meant it was a golden opportunity for Vivek to reap some of the new genetic resources.
“This was a good opportunity to use African germplasm, bring it into India and cross it to some Asia-adapted material,” he says.
Stored maize ears hanging in long bunches outside a house in China.
A key issue Vivek faced, however, was that most African maize varieties are white, and most Asian maize varieties are yellow. “You cannot directly deploy what you breed in Africa into Asia,” Vivek says. “Plus, there’s so much difference in the environments [between Africa and Asia] and maize is very responsive to its environment.”
The advances in marker-assisted breeding since the inception of GCP contributed significantly towards the success of Vivek’s team.
“In collaboration with GCP, IITA, Cornell University and Monsanto, CIMMYT has initiated the largest public sector MARS breeding approach in the world,” says Vivek.
The outcome is good: “We now have some early-generation, yellow, drought-tolerant inbred germplasm and lines suitable for Asia.
“GCP gave us a good start. We now need to expand and build on this,” says Vivek.
GCP’s supported work laid the foundation for other CIMMYT projects, such as the Affordable, Accessible, Asian Drought-Tolerant Maize project funded by the Syngenta Foundation for Sustainable Agriculture. This project is developing yet more germplasm with drought tolerance.
A better picture: GCP brightens maize research
Dickson Ligeyo’s worries of a stormy future for Kenya’s maize production have lifted over the 10 years of GCP. At the end of 2014, Kenya had two new varieties that were in the final stage of testing in the national performance trials before being released to farmers.
“There is a brighter picture for Kenya’s maize production since we have acquired acid-tolerant germplasm from Brazil, which we are using in our breeding programmes,” Dickson says.
In West Africa, researchers are also revelling in the opportunity they have been given to help enhance local yields in the face of a changing climate. “My institute benefited from GCP not only in terms of human resource development, but also in provision of some basic equipment for field phenotyping and some laboratory equipment,” says Allen Oppong in Ghana.
“Through the support of GCP, I was able to characterise maize landraces found in Ghana using the bulk fingerprinting technique. This work has been published and I think it’s useful information for maize breeding in Ghana – and possibly other parts of the world.”
The main challenge now for breeders, according to Allen, is getting the new varieties out to farmers: “Most people don’t like change. The new varieties are higher yielding, disease resistant, nutritious – all good qualities. But the challenge is demonstrating to farmers that these materials are better than what they have.”
This Kenyan farmer is very happy with his healthy maize crop, grown using an improved variety during a period of drought.
Certainly GCP has strengthened the capacity of researchers across Africa, Asia and Latin America, training researchers in maize breeding, data management, statistics, trial evaluations and phenotyping. The training has been geared so that scientists in developed countries can use genetic diversity and advanced plant science to improve crops for greater food security in the developing world.
Elliot Tembo, a maize breeder with the private sector in sub-Saharan Africa says: “As a breeder and a student, I have been exposed to new breeding tools through GCP. Before my involvement, I was literally blind in the use of molecular tools. Now, I am no longer relying only on pedigree data – which is not always reliable – to classify germplasm.”
Allen agrees: “GCP has had tremendous impact on my life as a researcher. The capacity-building programme supported my training in marker-assisted selection training at CIMMYT in Mexico. This training exposed me to modern techniques in plant breeding and genomics. Similarly, it built my confidence and work efficiency.”
There is no doubt that GCP research has brightened the picture for maize research and development where it is most needed: with researchers in developing countries where poor farmers and communities rely on maize as their staple food and main crop.
Rice plays a key role in global food security, particularly in Asia, where 90 percent of the world’s rice is grown and eaten. By 2050, Asia’s population is estimated to grow by one billion to 5.2 billion people, who will continue to depend on rice as their major staple food.
But with rising demand for rice has also come increasing salinity, droughts and other stresses, along with decreasing areas of land available for farming the crop.
Key ingredient in the rice research fest was GCP’s relationship with the International Rice Research Institute (IRRI), headquartered in The Philippines. GCP supported IRRI in its endeavours to use the latest molecular plant-breeding techniques, along with traditional plant-breeding tools, to develop rice crops better able to cope with various stresses and still be productive.
“These ‘super’ crops will revolutionise rice farming,” says IRRI Director General Robert Zeigler, who was also the first Director of GCP.
Climate change is one of the major threats facing rice production. As sea levels rise, salt water enters further up rivers with the high tides and affects rice production areas.
Each year in Bangladesh, during the boro rice season from November to May, salinity is so high that a white film of salt covers the country’s coastal paddy fields. For Bangladeshi farmers, this white colour is a warning sign that their land is ‘sick’. Around the world, Bangladesh, India, Myanmar and parts of Africa are most affected by increasing salinity.
Right from the beginning of GCP, salinity was a problem firmly on the rice research agenda.
Highly saline soils in India (left), and a close-up showing a surface crust of salt on afflicted soil (right).
Leading this research was IRRI plant physiologist Abdelbagi Ismail, who dreamed of the ‘super’ rice crop that could “tolerate salinity, drought and submergence”.
Abdelbagi has managed and overseen most of the progress made during the discovery of a major genetic region, or quantitative trait locus (QTL), associated with salinity tolerance and named Saltol.
Abdelbagi Ismail examines rice plants in the field in Bangladesh.
Its insertion into well-known rice varieties used by farmers in Bangladesh, Indonesia and The Philippines is part of the revolution taking place in rice research.
Abdelbagi says Saltol was mapped and markers were developed for its use in breeding more salt-tolerant rice varieties. Its salt-tolerance code is now being transferred into several varieties evaluated with IRRI partners in South and Southeast Asia.
“These projects also provided opportunities for both degree training and non-degree training to several of our partners in the countries involved,” he adds.
“Partnerships are crucial for us to build the capacity of the researchers in these countries and to ensure our outputs and outcomes reach the farmers that need them.
“All our partners benefitted from salt-tolerant varieties developed conventionally through this project, and they also provided pipelines for uptake and dissemination by farmers.”
Having developed new lines following the discovery of the Saltol QTL, Abdelbagi’s GCP-supported team trained plant breeders in country programmes to successfully breed for salt tolerance and other stresses.
In this way, Abdelbagi says, they are improving the capacity of researchers in developing countries to take up new breeding techniques, such as the use of molecular markers. “This can reduce the time it takes to breed new varieties, from six to ten years at the moment, down to two or three years,” he says.
This means that benefits to smallholder rice farmers struggling with salinity will happen sooner rather than later. And Abdelbagi credits GCP’s partnership approach, working directly with the countries in need, for the success so far.
“The salt-tolerant varieties are now being widely distributed,” he says. “Some of these varieties have doubled farmers’ productivity in affected areas.
“The work developed technologies of value to our needy farmers.
“We do believe this is the start of a second Green Revolution, especially for those who farm in less favourable areas and that missed this opportunity during the first Green Revolution.”
Partnership approach key to new rice gene for uptake of phosphorus
More than 60 percent of rainfed lowland rice is produced on poor and problem soils, including those that are naturally low in phosphorus. This is an essential for nutrient growing crops, but providing phosphorus through fertilisers is costly and unfeasible for many smallholder rice growers.
IRRI’s Sheryl Catausan prepares the roots of a rice plant for scanning as part of the work of the PSTOL1, phosphorus uptake research team at IRRI.
This problem was the focus of GCP’s rice phosphorus-uptake project led by IRRI molecular biologist Sigrid Heuer.
The project was enormously successful, with its discovery of the PSTOL1 (‘phosphorus starvation tolerance 1’) gene within the Pup1 locus, which was published in the prestigious journal Nature.
“We wanted it in Nature for a couple of reasons,” she says. “To raise awareness about phosphorus deficiency and phosphorus being a limited resource, especially in poorer countries; and to draw attention to how we do molecular breeding these days, which is a speedier, easier and more cost-effective approach to developing crops that have the potential to alleviate such problems.”
Following the PSTOL1 discovery, researchers are now working with developing-country researchers and extension agencies to help them understand how to breed local varieties of rice that can be grown in phosphorus-deficient soils. They are also collaborating with other crop breeders looking to breed similar maize, sorghum and wheat varieties.
Tobias Kretzschmar, a molecular biologist with IRRI, says that GCP’s partnership approach was the key to the project having an impact on the rice farmers who needed it most.
“For me, the collaborations that were forged through these inter-institutional projects made the difference,” he says.
Sigrid agrees: “GCP was always there supporting us and giving us confidence, even when we were not sure.”
Solving the insoluble: a gene for drought tolerance
Rice is a crop that not only needs water, but loves water. So developing a drought-tolerant rice variety is a quest to find a seemingly impossible gene.
However, GCP-supported researchers did just that: they solved the insoluble.
“They were very successful in terms of getting drought tolerance,” says Hei Leung of IRRI, who was GCP Subprogramme Leader for the Comparative Genomics Research Initiative between 2004 and 2007, and also a Principal Investigator for the Rice Research Initiative. “They’re now getting a 1.5 tonne rice yield advantage under water stress. I mean, that’s unheard of! This is a crop that needs water.
“This is one of GCP’s big success stories; that you can actually get drought tolerance is a seemingly impossible task for a water-loving rice plant.”
As Subprogramme Leader, Hei played a critical role in the creation, management, delivery and communication of a wide portfolio of research projects. He credits the nature of how GCP was set up for accelerating the breeding programme for drought-tolerant rice.
“The advantage of GCP is that it is run by a small group of people who can make fast decisions,” he says. “This means they can respond to the needs of researchers: ‘Okay, we are going to invest on that. We’re going to have a meeting on this’.
“The real advantage of GCP is its agility. Usually with other organisations you have new ideas and then have to slave away for a year to get the funding to implement them. But GCP was quick.”
Hei Leung (right) explains rice screening processes to a visiting scholar at IRRI.
IRRI and GCP deliver genetic resources to those who need them most
Tobias says one of the main objectives of GCP and IRRI is to make genetic stocks available to breeders, particularly in developing countries.
“Without that, IRRI would fail in its central mission to reduce hunger and poverty,” he says. “In order to achieve this mission, tight collaboration with our agricultural and extension partners in other countries is the key.”
In fact, this idea of protecting the future through genetic material was influential in the choice of GCP as a CGIAR Challenge Program name. Hei, who was involved from the start with GCP, talks about the meeting in a Rome pizzeria where participants came up with the name: “When we talk about ‘generation’, we are really talking about the work we do with genetic diversity; it is about the future generation,” he says.
Part of that future generation is about sharing genetic resources or stocks, but first the genetic diversity of such stocks needs to be characterised. Hei remembers that one of the first GCP projects in 2004 brought together researchers in various countries to characterise the genetic diversity of various crops, including rice.
“But everyone was using different genotyping platforms and markers, and the technology back then was not what we have now,” he recalls.
“So we spent a lot of resources getting poor quality data. In a sense, it was a failure. On the other hand, it was also a success because we alerted people to the importance of characterising diversity in every single crop. The whole concept of getting all the national partners doing genetic resource characterisation is a good one.
“We have evolved the technology together over the last 10 years of GCP. Now the country partners feel enabled. They are not scared of the technology.”
Abdelbagi agrees that characterising genetic diversity is essential, and adds that making such genetic stocks readily available to breeders is also vital.
“This has not been an issue before,” he explains. “With the new regulations of germplasm control and intellectual property issues, it became extremely difficult to exchange germplasm with some countries. One important lesson we learnt was to engage in direct discussion with our partners in efforts to influence their policies and guidelines to allow essential exchange of genetic stocks and breeding material, at least at the regional level.”
Abdelbagi adds that another big challenge has been to provide country partners with materials and DNA markers for marker-assisted selection programmes and to make sure these were properly used in breeding programmes.
“We solved this by hosting a workshop at IRRI and with continued visits with GCP collaborators,” he says.
Rice terraces in Ifugao Province in The Philippines.
‘Super’ crops: something ‘magic’ happens
Hei says that the GCP project he’s most passionate about, and one that leaves a lasting legacy, is the development of multiparent advanced generation intercross (MAGIC) populations, which will potentially yield lines that are tolerant to environmental stresses, grow well in poor soils and produce better quality grain.
“MAGIC populations will leave behind a very good resource towards improving different crop species,” says Hei. “I’m sure that they will expand on their own.”
GCP funded the development of four kinds of rice populations, including indica MAGIC, which is the most advanced of the MAGIC populations developed so far. These populations contain multiple desirable traits, including: blast and bacterial blight-resistance, salinity and submergence tolerance and grain quality.
New generation of researchers working on a new rice revolution
Rice farmer in her field in Rwanda.
Robert Zeigler’s dream of a new rice Green Revolution with ‘super crops’ is coming true, thanks in part to GCP’s focus on combining cutting-edge molecular plant-breeding techniques with conventional plant breeding.
“With all this going for us, the second Green Revolution means we can meet the great challenges ahead with unprecedented efforts that will result in unparalleled impacts,” he says. “This will range from mining the rice genomes for needed traits to developing climate change-ready rice.”
IRRI researchers like Abdelbagi agree that new plant-breeding techniques, such as those fostered by GCP, are making ‘super’ crops more likely: “I’m committed to understand how plants can be manipulated to adapt to, and better tolerate, extreme environmental stresses, which seems more feasible today than it has ever been before.
“GCP-supported work has provided mechanisms for developing varieties with multiple stress tolerances, besides the improvements in yield and quality.”
And for Hei, GCP’s 10-year legacy is not just about the technology but also about the people.
“Over ten years you have three generations of PhD students,” he says. “Many people became a ‘new generation’ scientist through this programme. Many people have benefitted. GCP is one of a kind. I’m just in love with it.”
GCP-supported researchers aimed high: they wanted to contribute to food security in the developing world by using the latest advances in crop science and plant breeding.
And with the lives of half of the world’s population directly reliant on their own agriculture, there is a lot at stake. Land degradation, salinity, pollution and excessive fertiliser use are just some of the challenges.
Rice is one of the most critical crops worldwide
Amelia Henry, drought physiology group leader at the International Rice Research Institute (IRRI), explains why rice was such a critical crop for GCP research. She says rice is grown in a diverse set of environmental settings, often characterised by severe flooding, poor soils and disease.
Cycling through rice fields in Odisha, India.
In Asia, 40 percent of rice is produced in rainfed systems with little or no water control or protection from floods and droughts – meaning rice plants are usually faced with too much or too little water, and rarely get just enough. In addition, 60 percent (29 million hectares) of the rainfed lowland rice is produced on poor and problem soils, including those that are naturally low in phosphorus.
Phosphorus deficiency and aluminium toxicity are two of the most widespread environmental causes of poor crop productivity in acidic soils, where high acid levels upset the balance of available nutrients. And drought makes these problems even worse.
Phosphorus is essential for growing crops. Its commercial use in fertilisers is due to the need to replace the phosphorus that plants have extracted from the soil as they grow. Soils lacking phosphorus are an especially big problem in Africa, and the continent is a major user of phosphate fertilisers. However, inappropriate use of fertilisers can, ironically, acidify soil further, since excess nitrogen fertiliser decreases soil pH.
Meanwhile, high levels of aluminium in soil cause damage to roots and impair crop growth, reducing their uptake both of nutrients like phosphorus and of water – making plants more vulnerable to drought. Aluminium toxicity is a major limitation on crop production for more than 30 percent of farmland in Southeast Asia and South America and approximately 20 percent in East Asia, sub-Saharan Africa and North America.
The challenge today is to tap into the genetic codes of key crops such as rice and wheat to feed a growing global population. Science plays a crucial role in identifying genes for traits that help plants tolerate more difficult environmental conditions, and producing crop varieties that contain these genes.
Plant biologists are already developing new rice lines that produce higher yields in the face of reduced water, increasingly scant fertiliser as costs rise, and unproductive soils. However, ‘super’ crops are needed that can combine these qualities and withstand climate changes such as increasing temperatures and reduced rainfall in a century when the world’s population is estimated to reach nearly 10 billion people by 2050.
Bringing the best scientific minds to improve rice varieties
Ambitious in concept, the GCP research focussed on bringing together experts to work on these critical problems of rice production for some of the world’s poorest farmers.
The programme was rolled out in two phases that sought to explore the genetic diversity of key crops and use the most important genes for valuable traits, such as Sigrid’s discovery made in a rice variety that is tolerant of phosphorus-poor soils. Each phase involved dedicated teams in partner countries.
GCP Principal Investigator Hei Leung, from IRRI, says GCP is unique, one of kind: “I love it.” He says GCP has enabled rice researchers and breeders to embrace cutting-edge science through partnerships focussed on improving crop yields in areas previously deemed unproductive.
Hei says GCP wanted to target research during its second phase on those crops that most poor people depend upon. “We wanted to have a programme that is what we call ‘pro-poor’, meaning the majority of the world’s people depends on those crops,” he says.
Rice is the ‘chosen one’ of GCP’s cereal crop research and development, with the biggest slice of GCP’s research activities dedicated to this, the most widely consumed staple food.
It is crucial to increase rice supplies by applying research and development such as that carried out by GCP researchers over the past 10 years, Hei says.
Relying on rice’s small genome in the hunt for drought-tolerance genes
Researchers had been trying to map the genomes of key cereal crops for over two decades. Rice’s genome was mapped in 2004, just as GCP started.
Rice has a relatively small genome, one-sixth the size of the maize genome and 40 times smaller than the wheat genome. This makes it a useful ‘model’ crop for researchers to compare with other crops.
“People like to compare with rice because wheat and maize have very big genomes, and they don’t have the resources,” explains Hei.
After the rice genome had been sequenced, the next step was to focus down to a more detailed level: the individual genes that give rice plants traits such as drought tolerance. Identifying useful genes, and markers that act as genetic ‘tags’ to point them out, gives scientists an efficient way to choose which plants to use in breeding.
One of GCP’s Principal Investigators for rice was Marie-Noëlle Ndjiondjop, a senior molecular scientist with the Africa Rice Center.
“Rice is becoming a very important crop in Africa,” she says. “Production has been reduced by a lot of constraints, and drought is one of the most important constraints that we face in Africa.”
Meet Marie-Noëlle below (or on YouTube), in our series of Q&A videos on rice research in Africa.
Marie-Noëlle’s team recognised that drought tolerance was likely to be a complex trait in rice, involving many genes, due to the mix of physiological, genetic and environmental components that affect how well a plant can tolerate drought conditions. To help discover the rice varieties likely to have improved drought tolerance, Marie-Noëlle’s team used an innovative approach known as bi-parental marker-assisted recurrent selection (MARS).
“With such a complex trait, you really need to have all the tools and infrastructure necessary; through GCP we were able to buy the necessary equipment and put in the infrastructure needed to find and test the drought trait in rice lines.
“By using the MARS approach we identified the genetic regions associated with drought and are moving towards developing new rice lines that the African breeder and farmer will be using in the next decade to grow crops that are better able to withstand drought conditions.”
Likewise, Amelia Henry’s IRRI team also developed drought-tolerant lines, particularly for drought-prone areas of South Asia. She says many of the promising deep-rooted or generally drought-tolerant varieties identified in the early decades after IRRI’s foundation in 1960 are still used today as ‘drought donors’.
“Since the strength of our project was the compilation of results from many different sites, this work couldn’t have been done without the GCP partners,” she says. “They taught me a lot about how rice grows in different countries and what problems rice farmers face.”
Hei agrees that GCP partnerships have been crucial, including in the successful breeding of rice with drought tolerance: “They’re getting a 1.5-tonne rice yield advantage under water stress. I mean, that’s unheard of! This is a crop that needs water.”
A rice farmer in Rwanda.
But the researchers could not rest with just one of rice’s problems solved.
Hei says GCP’s initial focus on drought was a good one but then, “I remember saying, ‘We cannot just go for drought. Rice, like all crops, needs packages of traits’.”
He knows that drought is just one problem facing rice farmers, noting “this broadened our research portfolio to include seeking to breed rice varieties with traits of tolerance to aluminium toxicity, salt and poor soils.”
The scope widens: phosphorus-hungry rice and a huge success
Sigrid Heuer was in The Philippines working for IRRI when she became involved in the ground-breaking phosphorus-uptake project for rice.
She took over the project being headed by Matthias Wissuwa. Much earlier, Matthias had noted that Kasalath – a traditional northern Indian rice variety that grew successfully in low-phosphorus soil – must contain advantageous genes. His postdoctoral supervisor, Noriharu Ae, thought that longer roots were likely to be the secret to some rice varieties being able to tolerate phosphorus-deficient soils.
Screening for phosphorus-efficient rice, able to make the best of low levels of available phosphorus, on an IRRI experimental plot in The Philippines. Some types of rice have visibly done much better than others.
Sigrid Heuer used her background in molecular breeding to take up the challenge with GCP to find the genes responsible for the Kasalath variety’s long roots.
“I spent years looking for the gene,” Sigrid says. “It was like trying to find a needle in a haystack; the genomic region where the gene is located is very complex.
“We had little biogenomics support at the time and I had three jobs and two kids; I was spending all my nights trying to find this gene.”
Sigrid Heuer in the field at IRRI.
But one day, Sigrid’s postdoctoral student Rico Gamuyao excitedly called her downstairs to the transgenic greenhouses. “Rico had used transgenic plants to see whether this gene had any effect. He was digging out plants from experimental pods.”
Sigrid says that moment in the Manila labs was the turning point for the project’s researchers.
Matthias’ team had previously identified a genomic region, or locus, named Pup1 (‘phosphorus uptake 1’) that was linked to phosphorus uptake in lines of traditional rice growing in poor soils. However, its functional mechanism remained elusive until the breakthrough GCP-funded project sequenced the locus, showing the presence of a Pup1-specific protein kinase gene, which was named PSTOL1 (‘phosphorus starvation tolerance 1’). The discovery was reported in the prestigious scientific journal Nature on 23 August 2012 and picked up by media around the world.
The gene instructs the plant to grow larger and longer roots, increasing its surface area – which Sigrid compares to having a bigger sponge to absorb more water and nutrients in the soil.
“Plants growing longer roots have more uptake of phosphorus – and PSTOL1 is responsible for this.
“GCP was always there, supporting us and giving us confidence, even when we weren’t sure we were going to succeed,” she recalls. “They really wanted us to succeed, so, financially and from a motivational point of view, this gave us more enthusiasm.”
She adds, jokingly, “With so many people having expectations about the project, it was better not to disappoint.”
For some insight straight from the source, listen to Matthias in our podcosts below. In these two bitesized chunks of wisdom he discusses the importance of phosphorus deficiency and of incorporating PSTOL1 into national breeding programmes; his work in Africa and the possibility of uncovering an African ‘Pup2’; what the PSTOL1 discovery has meant for him; and the essential contribution of international partnerships and GCP’s support.
Members of the IRRI PSTOL1, phosphorus uptake research team chat in the field in 2012. From left to right they are are: Sigrid Heuer, Cheryl Dalid, Rico Gamuyao, Matthias Wissuwa and Joong Hyoun Chin.
Phosphorus-uptake gene not all it seemed – an imposter?
But PSTOL1 was definitely not what it seemed. “It was identified under phosphorus-deficient conditions and the original screen was set up for that,” says Sigrid.
Researchers eventually discovered that Pup1 and the PSTOL1 gene within it were not really all about phosphorus at all: “It turns out it is actually a root-growth gene, which just happens to enhance uptake of phosphorus and other nutrients such as nitrogen and potassium.
“The result is big root growth and maintenance of that growth under stress. If you have improved root growth, there is more access to soil resources, as a plant can explore more soil area with more root fingers.”
Her team showed that overexpression of PSTOL1 gene significantly improves grain yield in varieties growing in phosphorus-deficient soil – by up to 60 percent compared to rice varieties that did not have the gene.
In field tests in Indonesia and The Philippines, rice with the PSTOL1 gene produced about 20 percent more grain than rice without the gene. This is important in countries where rice is grown in poor soils.
A farmer harvests rice in South Sulawesi, Indonesia.
Sigrid, now based in Adelaide at the Australian Centre for Plant Functional Genomics, says the introduction of the new gene into locally adapted rice varieties in different locations across Asia and Africa is expected to boost productivity under low-phosphorus conditions.
“The ultimate measure for these kinds of projects is whether a gene works in different environments. I think we have a lot of evidence that says it does,” she says.
The discovery of PSTOL1 promises to improve the food security of rice farmers on phosphorus-deficient land though assisting them to grow more rice and earn more.
Titbits of further research successes: aluminium tolerance and MAGIC genes
Drought, low-phosphorus soils, aluminium toxicity, diseases, acid soils, climate change… the list seems never-ending for challenges to growing rice. Apart from the successes with drought and phosphorus that GCP scientists achieved, there was to be much more in the works from other GCP researchers.
In Phase II, they worked towards breeding aluminium-tolerant sorghum lines for sub-Saharan Africa, as well as applying what they learnt to discover similar genes in rice and maize.
Hei Leung says GCP leaves a lasting legacy in the development of multiparent advanced generation intercross (MAGIC) populations. These help breeders to identify valuable genes, and from among the populations they can also select lines to use in breeding that have favourable traits, such as being tolerant to environmental stresses, having an ability to grow well in poor soils or being able to produce better quality grain.
“MAGIC populations will leave behind a very good resource towards improving different crop species,” says Hei. “I’m sure that they will expand on their own.”
GCP funded the development of four different MAGIC populations for rice, including both indica and japonica types. And the idea of developing MAGIC populations has spread to other crops, including chickpeas, cowpeas and sorghum.
Meeting the challenges and delivering outcomes to farmers
But with success come the frustrations of getting there, according to Nourollah Ahmadi, GCP Product Delivery Coordinator for rice across Africa. “This is because things are not always going as well as you want.”
Project Delivery Coordinators monitor projects first-hand, conducting on-site visits, advising project leaders and partners and helping them implement delivery plans.
“One of the problems was the overall level of basic education of people who were involved in the project,” Nourollah says.
Rice cultivation in Mali is on the rise.
His work with GCP has opened up new prospects for some of the poorest farmers in the world: “For five years, I have been coordinating one of the rice initiatives implemented by the Africa Rice Center and involving three African countries.” These are Burkina Faso, Mali and Nigeria.
He says GCP has brought much-needed expertise and technical skills to countries which can now use genetic insights to produce improved crops tolerant of drought conditions and poor soils and resistant to diseases. Using new molecular-breeding techniques has provided a more effective way to move forward, still firmly focussed on helping the world’s poorest farmers achieve food security.
“We don’t change direction, we change tools – sometimes you have a bicycle, sometimes you have a car,” Nourollah says.
Hei agrees there have been challenges: “It’s been a bumpy road to get to this point. But the whole concept of getting all the national partners doing genetic resource characterisation is a very good one.
Right now they are enabled; they are not scared about the technology. They can apply it.”
Sigrid says applied research is judged on two scales: “One is the publications and science you’re doing. The other is whether the work has any impact in the field, whether it works in the field. Bringing these two together is sometimes a challenge.”
GCP has managed to meet both challenges. New crop varieties have been released to farmers, and more than 450 scientifically reviewed papers have been published since 2004.
Building on the rice success story and leaving a lasting legacy
The work that GCP-supported researchers have done for rice is also being used in other crops. For example, researchers used comparative genomics to determine if genes the same as or similar to those found in rice are present and operating in the same manner in sorghum and maize.
The GCP team found sorghum and maize varieties that contained genes, similar to rice’s PSTOL1, that also confer tolerance of phosphorus-deficient soil with an enhanced root system. They were then able to develop markers to help breeders in Brazil and Africa identify phosphorus-efficient lines.
The knowledge that GCP-supported rice researchers have generated is shared through communities of practice, through websites, publications, research meetings and the Integrated Breeding Platform.
As Amelia Henry notes, GCP’s achievements will be defined by “the spirit of dedication to openness with research data, results and germplasm and giving credit and support to partners in developing countries.” The work in rice in many ways exemplifies GCP’s collaborative approach, commitment to capacity building and deeply held belief that together we can go so much further in helping farmers.
Unlocking genetic diversity in crops for the resource-poor was at the heart of GCP’s mission, which in 2003 promised ‘a new, unique public platform for accessing and developing new genetic resources using new molecular technologies and traditional means’.
Certainly for poor rice farmers in Asia and Africa, the work that GCP has supported in applying the latest molecular-breeding techniques will lead to rice varieties that will help them produce better crops on poor soils in a changing climate.