Generation Challenge Programme
GCP website
Integrated Breeding
IBP website
GCP Blog
GCP blog
  Connect with us GCP on Facebook GCP on Twitter GCP on LinkedIn Subscribe to GCP Newsletter Subscribe to GCP RSS feeds
Oct 262015
 

 

Photo: HK Tang/Flickr (Creative Commons)

An Indian patchwork of rice and maize fields.

“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.

Leon was the Product Delivery Coordinator for the Comparative Genomics Research Initiative of the CGIAR Generation Challenge Programme (GCP). The Programme set out in 2004 to advance plant genetics in a bid to provide sustainable food-security solutions.

Between 2004 and 2014, GCP invested in projects to clone two genes and deploy them in elite local varieties. The first gene, SbMATE, encodes aluminium tolerance traits in sorghum; the work was a collaborative effort led by the Brazilian Corporation of Agricultural Research (EMBRAPA) and Cornell University, and gave rise to locally-adapted sorghum varieties released in South America and Africa. The second gene, PSTOL1, produces traits that improve phosphorus uptake in rice. This research was a collaboration between the Japan International Research Center for Agricultural Sciences (JIRCAS) and the International Rice Research Institute (IRRI). PSTOL1 has now been extensively deployed in Asian rice varieties.

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.

Photo: S Kilungu/CCAFS

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.

Photo: ICRISAT

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

Photo: IRRI

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.”

Photo: IRRI

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.

Photo: S Kilungu/CCAFS

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.”

More links

Photo: CSISA

A rice farmer in Bihar, India.

Oct 122015
 

 

Photo: One Acre Fund/Flickr (Creative Commons)

A Kenyan farmer harvesting her maize.

“The map of Kenya’s maize-growing regions mirrors the map of the nation’s acid soils.”

So says Dickson Ligeyo, senior research officer at the Kenya Agricultural and Livestock Research Organisation (KALRO; formerly the Kenya Agricultural Research Institute, or KARI), who believes this paints a sombre picture for his country’s maize farmers.

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.

Photo: Allison Mickel/Flickr (Creative Commons)

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.

 By weight, more maize is produced each year than any other grain: global production is more than 850 million tonnes. Maize production is increasing at twice the annual rate of rice and three times that of wheat. In 2020, demand for maize in developing countries alone is expected to exceed 500 million tonnes and will surpass the demand for both rice and wheat.  This projected rapid increase in demand is mainly because maize is the grain of choice to feed animals being reared for meet – but it is placing strain on the supply of maize for poor human consumers. Demand for maize as feed for poultry and pigs is growing, particularly in East and Southeast Asia, as an ever-increasing number of people in Asia consume meat. In some areas of Asia, maize is already displacing sorghum and rice. Acreage allocated to maize production in South and Southeast Asia has been expanding by 2.2 percent annually since 2001. In its processed form, maize is also used for biofuel (ethanol), and the starch and sugars from maize end up in beer, ice cream, syrup, shoe polish, glue, fireworks, ink, batteries, mustard, cosmetics, aspirin and paint.

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.

Photo: A Wangalachi/CIMMYT

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.

Scientists join hands to unravel maize complexity

Scientists from the International Maize and Wheat Improvement Center (CIMMYT) and the International Crops Research Institute for the Semi-Arid Tropics (ICRISAT) got their heads together between 2005 and 2008 to itemise what maize stocks were available.

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.

Photo: X Fonseca/CIMMYT

Maize diversity.

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.

Photo: N Palmer/CIAT

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.

Photo: L Kochian

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.”

The intensity of GCP-supported maize research shifted up a gear in 2007, after the team led by Jurandir Magalhães, research scientist in molecular genetics and genomics of maize and sorghum at EMBRAPA, used positional cloning to identify the major sorghum aluminium tolerance gene SbMATE responsible for the AltSB aluminium tolerance locus. The team comprised researchers from EMBRAPA, Cornell, the Japan International Research Center for Agricultural Sciences (JIRCAS) and Moi University in Kenya.

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.

Photos: 1 – V Alves ; 2 – F Mendes; both edited by C Guimarães

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.

Marker-assisted selection helps breeders like Sam Gudu more quickly develop plants that have desirable genes. When two plants are sexually crossed, both positive and negative traits are inherited. The ongoing process of selecting plants with more desirable traits and crossing them with other plants to transfer and combine such traits takes many years using conventional breeding techniques, as each generation of plants must be grown to maturity and phenotyped – that is, the observable characteristics of the plants must be measured to determine which plants might contain genes for valuable traits.   By using molecular markers that are known to be linked to useful genes such as ZmMATE1, breeders can easily test plant materials to see whether or not these genes are present. This helps them to select the best parent plants to use in their crosses, and accurately identify which of the progeny have inherited the gene or genes in question without having to grow them all to maturity. Marker-assisted selection therefore reduces the number of years it takes to breed plant varieties with desired traits.

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.

Photo: N Palmer/CIAT

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.”

Photo: N Palmer/CIAT

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.

Photo: D Mowbray/CIMMYT

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.

Photo: A Erlangga/CIFOR

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.

Photo: E Phipps/CIMMYT

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.”

Photo: CIMMYT

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.

More links

Photo: N Palmer/CIAT

A farmer displays maize harvested on his farm in Laos.

Dec 052014
 

It’s a cruel feature of some of the most populous areas of the world, particularly in the tropics and subtropics: acid soils. They cover a third of the world’s total land area – including significant swathes of Africa, Asia and Latin America – and 60 percent of land we could use for growing food. Today around 30 percent of all arable land reaches levels of acidity that are toxic to crops.

Soil acidity occurs naturally in higher rainfall areas and varies according to the landscape and soil. But we also make the problem worse through intensive agricultural practices. The main cause of soil acidification is the overuse of nitrogen fertilisers, which farmers apply to crops to increase production. Ironically, the inefficient use of nitrogen fertiliser can instead make matters worse by decreasing the soil pH.

60 percent of the world’s potential crop-growing land is highly acidic. Map courtesy of Leon Kochian.

60 percent of the world’s potential crop-growing land is highly acidic. Map courtesy of Leon Kochian.

Acidity prevents crops from accessing the right balance of nutrients in the soil, limiting farmers’ yields. Its negative effect on world yield is second only to drought and is particularly hard felt by subsistent and smallholder farmers who cannot afford to correct soil pH using calcium-rich lime. As a result, these farmers are forced to grow less profitable, acid-tolerant crops like millet, or suffer huge yield losses when growing more popular cereal crops like wheat, rice or maize.

“In Kenya, acidic soils cover almost 90 percent of the maize-growing areas and can reduce yields by almost 60 percent,” says Samuel Gudu, Professor and Deputy Vice-Chancellor (Planning & Development) at Moi University in Kenya. “Farmers know that the soil affects their yields, but they still grow maize because it is so popular.”

As is true in many other sub-Saharan countries, maize is a staple of the Kenyan diet: the average Kenyan consumes 98 kilograms of it each year. But maize prices in Kenya are among the highest in Africa, which directly affects the poorest quarter of the population, who spend 28 percent of their income on the crop.

“Yield losses play a big part in this economic imbalance and are why we need affordable agronomic options to help our farmers improve yields,” says Samuel, who was a Principal Investigator of a GCP comparative genomics project which sought to provide some of these options.

A Kenyan farmer prepares her maize plot for planting. Acid soils cover almost 90 percent of Kenya’s maize-growing area, and can more than halve yields.

A Kenyan farmer prepares her maize plot for planting. Acid soils cover almost 90 percent of Kenya’s maize-growing area, and can more than halve yields.

Aluminium toxicity and phosphorus deficiency: Public enemies number one and two in the fight against acidic soils

Between 2004 and 2014, crop researchers and plant breeders across five continents collaborated on several GCP projects to develop local varieties of maize, rice and sorghum that can withstand phosphorus deficiency and aluminium toxicity – two of the most widespread constraints leading to poor crop productivity in acidic soils.

Aluminium toxicity is the primary limitation on crop production for more than 30 percent of farmland in Southeast Asia and Latin America and approximately 20 percent in East Asia, sub-Saharan Africa and North America. Aluminium becomes more soluble in acid souls, creating a toxic glut of aluminium ions that damage roots and impair their growth and function. This results in reduced nutrient and water uptake, which in turn depresses yield.

Phosphorus deficiency is the next biggest soil deficiency after nitrogen to limit plant production. In acid soils, phosphorus is stuck (fixed) in forms that plants cannot take up. All plants need phosphorus to survive and thrive; it is a key element in plant metabolism, root growth, maturity and yield. Plants deficient in phosphorus are often stunted.

In a double whammy, the damage that aluminium toxicity causes to roots means that plants cannot efficiently access native soil phosphorus or even added phosphorus fertiliser – and adding phosphorus is an option that is rapidly becoming less viable.

“The world is running out of phosphorus as quickly as it is running out of oil,” says Leon Kochian, a Professor in the Departments of Plant Biology and Crop and Soil Science at Cornell University in the USA. “This is making its application a more expensive and less sustainable option for all farmers wanting to improve yields on acidic soils.” Indeed, the price of rock phosphate has more than doubled since 2007.

For 30 years, Leon has combined lecturing and supervising duties at Cornell University and the United States Department of Agriculture with his scientific quest to understand the genetic and physiological mechanisms that allow some cereals to tolerate acidic soils while others wither. And for the last 10 years, he has played an important leading role in GCP’s effort to develop new, higher yielding varieties of maize, rice and sorghum that tolerate acidic soils.

GCP builds on past crop breeding successes

The rationale behind GCP’s efforts stems from two independent and concurrent projects, which had been flourishing on different sides of the Pacific well before GCP was created.

One of those projects was co-led by Leon at Cornell University in collaboration with a previous PhD student of his, Jurandir Magalhães, at the Brazilian Corporation of Agricultural Research (EMBRAPA) Maize & Sorghum research centre.

Working on the understanding that the cells in grasses like barley and wheat use ‘membrane transporters’ to insulate themselves against excessive subsoil aluminium, Leon and Jurandir searched for a similar transporter in the cells of sorghum varieties that were known to tolerate aluminium.

“In wheat, when aluminium levels are high, these membrane transporters prompt organic acid release from the tip of the root,” explains Jurandir. “The organic acid binds with the aluminium ion, preventing it from entering the root.” Jurandir’s team found that in certain sorghum varieties, the gene SbMATE encodes a specialised organic acid transport protein, which stimulates the release of citric acid. They cloned the gene and found it was very active in aluminium-tolerant sorghum varieties. They also discovered that the activity of SbMATE increases the longer the plant is exposed to high levels of aluminium.

The rice variety on the left (IR-74) has the the gene locus Pup1, conferring phosphorus-efficient longer roots, while the rice on the right does not.

The rice variety on the left (IR-74) has the the gene locus Pup1, conferring phosphorus-efficient longer roots, while that on the right does not.

The other project, co-led by Matthias Wissuwa at Japan International Research Centre for Agricultural Sciences (JIRCAS) and Sigrid Heur at the International Rice Research Institute (IRRI) in The Philippines, was looking for genes that could improve rice yields in phosphorus-deficient soils. They had already identified a gene locus (a section of the genome containing a collection of genes) that produced a protein which allowed rice varieties with to grow successfully in low-phosphorous conditions. The locus was termed ‘phosphorus uptake 1’ or Pup1 for short. With GCP support, the team were able to make the breakthrough of discovering the protein kinase gene responsible, PSTOL1 (‘phosphorus starvation tolerance 1’), and understanding its mechanism.

“In phosphorus-poor soils, this protein instructs the plant to grow larger, longer roots, which are able to forage through more soil to absorb and store more nutrients,” explains Sigrid, a plant geneticist at IRRI and a GCP Principal Investigator. “By having a larger root surface area, plants can explore a greater area in the soil and find more phosphorus than usual. It’s like having a larger sponge to absorb more water.”

Screening for phosphorus-efficient rice, able to make the best of low levels of available phosphorus, on an International Rice Research Institute (IRRI) experimental plot in the Philippines. Some types of rice have visibly done much better than others.

Screening for phosphorus-efficient rice, able to make the best of low levels of available phosphorus, on an International Rice Research Institute (IRRI) experimental plot in the Philippines. Some types of rice have visibly done much better than others.

Leon clarifies that both projects were fairly advanced before they became part of the GCP fold. “Our team had already identified the gene SbMATE and were in the process of cloning it for breeding purposes. The IRRI and JIRCAS team had also identified Pup1 and were in the process of identifying and cloning the gene.”

The purpose of cloning these genes was to create molecular markers to help breeders identify whether the genes were present in the varieties they were working with. As an analogy, think of ‘reading’ a plant’s genome as you would read a story: the story’s words are the plant’s genes, and a molecular marker works as a text highlighter. Different markers can highlight or tag different keywords in the story. Tagging the location of beneficial genes in the DNA of plant genomes allows scientists to see which of the plants or seeds they are interested in – perhaps only a few out of hundreds or thousands – contain these genes. This forms the basis of marker-assisted breeding, which can help plant breeders halve the time it takes them to breed new high-yielding varieties for acidic soil conditions.

Leon says that GCP provided both projects with the opportunity to validate their discoveries and to use what they had found to develop new aluminium-tolerant sorghum varieties and phosphorus-efficient rice varieties for farmers. But it’s what happened next that made this GCP initiative unique.

Finding the best genes within the crop family

Sorghum, rice, maize and wheat are all part of the Poaceae (true grasses) family, evolving from a common grass ancestor 65 million years ago. Over this time, they have become very different from each other. However, at the genetic level they still have a lot in common.

Over the last 20 years, genetic researchers all over the world have been mapping these cereals’ genomes. These maps are now being used by geneticists and plant breeders to identify similarities and differences between the genes of different cereal species. This process is termed ‘comparative genomics’ and was a fundamental research theme for GCP during its second phase (2008–2014).

“The objective during GCP Phase I (2004-2007) was to study the genomes of important crops and identify genes conferring resistance or tolerance to various stresses, such as drought,” says Rajeev Varshney, Director of the Center of Excellence in Genomics at the International Crops Research Institute for the Semi-Arid Tropics (ICRISAT). “This research was long and intensive, but it set a firm foundation for the work in GCP’s second phase, which sought to use what we have learnt in the laboratory and apply it to breed better varieties of crops.”

Rajeev oversaw GCP’s comparative genomics research projects on aluminium tolerance and phosphorus deficiency in sorghum, maize and rice, as part of his GCP role as Leader of the Comparative and Applied Genomics Research Theme.

“The idea behind the sorghum, maize and rice initiative was to use the discoveries we had independently made in sorghum and rice to see if we could find the same genes in the other crop,” explains Rajeev. “In other words, we wanted to see if we could find PSTOL1 in sorghum and SbMATE in rice.”

Working together through a number of comparative genomics projects, the researchers were highly successful in reaching this goal, discovering valuable sister genes and beginning to introduce them into new improved crop varieties for farmers.

Extending research in sorghum and rice to maize

Researchers at Cornell and EMBRAPA had already been using similar comparative techniques to look for SbMATE in maize because of its close familial connection to sorghum. This research was overseen by Leon and another EMBRAPA researcher, Claudia Guimarães.

“We used the knowledge that Jurandir and Leon’s SbMATE project produced to prove that we had a major aluminium-tolerance gene,” reflects Claudia.

The SbMATE gene in sorghum explains about 80 percent of its aluminium tolerance, but Claudia says that in maize it explains only about 20 per cent, making it harder for researchers to find without a little help knowing what to look for. “So we had to dig a little deeper for other similar genes that confer aluminium tolerance, and we found ZmMATE.”

Maize trials in the field at EMBRAPA. The maize plants on the left are aluminium-tolerant while those on the right are not.

Maize trials in the field at EMBRAPA. The maize plants on the left are aluminium-tolerant while those on the right are not.

ZmMATE1 has a similar genetic sequence to SbMATE and encodes a similar protein membrane transporter that releases citric acid from the roots. Just as in sorghum, citric acid binds to aluminium in the soil, making it difficult for it to enter plant roots. The team have also discovered related gene ZmMATE2, which also encodes a transporter protein, but appears to confer aluminium tolerance via a different mechanism, as yet unclear.

Claudia has developed a number of molecular markers for ZmMATE, which have been successfully used by breeders at EMBRAPA as well as by African partners in Niger and Kenya, such as Samuel Gudu, to identify maize breeding lines that have the gene.

“We used aluminium-tolerant maize varieties sourced locally and from Brazil to develop a range of potential new varieties,” says Samuel. “The goal is to develop varieties that are suited to our environment and not too dissimilar to varieties that Kenyan farmers like to grow, except they have a higher tolerance to aluminium toxicity.”

Left to right (foreground): Leon Kochian, Jurandir Magalhães and Samuel Gudu examine crosses between Kenyan and Brazilian maize, at the Kenya Agricultural Research Institute (KARI), Kitale, in May 2010.

Left to right (foreground): Leon Kochian, Jurandir Magalhães and Samuel Gudu examine crosses between Kenyan and Brazilian maize, at the Kenya Agricultural Research Institute (KARI), Kitale, in May 2010.

Involving farmers in the crop breeding process is an important part of such programs being successful, explains Samuel. “They help us identify maize varieties that they have observed have higher tolerance to acidic soils. We also try to incorporate other features that they want, such as disease resistance and higher yield. By incorporating their feedback into the breeding process they are more likely to grow any new varieties, as they have played a part in their development.”

Samuel says they have developed some local aluminium-tolerant varieties, which rank among the best for aluminium tolerance. Interestingly, these varieties seem to have a different aluminium-tolerance mechanism to the Brazilian varieties.

“From the work Samuel has done, we’ve possibly identified a novel source of aluminium tolerance in Kenyan maize varieties,” says Claudia. “We are now working together with Leon to identify the genes that are conferring this tolerance so we can develop markers to help Kenyan maize breeders also identify these varieties more efficiently.”

To help in the process, Samuel and his team are developing single-cross hybrids with a combination of both the novel Kenyan sources of aluminium tolerance and ZmMATE from Brazil, which will be even more tolerant to acidic soils.

Breeding for multiple stresses is a step-by-step process

Suradiyo, a farmer from Bojong Village near Yogyakarta, Indonesia, harvests rice.

Suradiyo, a farmer from Bojong Village near Yogyakarta, Indonesia, harvests rice.

In Asia, about 60 percent of rainfed rice is grown on soils that are affected by multiple stresses. These typically include phosphorus deficiency as well as aluminium toxicity, salinity and drought.

These stresses are particularly hard felt in Indonesia, which is the world’s third-largest rice producer. Joko Prasetiyono is a molecular rice breeder at the Indonesian Center for Agricultural Biotechnology and Genetic Resources Research and Development (ICABIOGRAD). His team have been collaborating with IRRI and JIRCAS for many years and contributed to validating the effect of Pup1 by embedding it into three popular local rice varieties – Dodokan, Situ Bagendit and Batur – which were then able to tolerate phosphorus-deficient conditions.

“The aim [with GCP research] was to breed varieties identical to those that farmers already know and trust, except that they have PSTOL1 and an improved ability to take up soil phosphorus,” says Joko.

Joko says that these varieties – which will be available in one to two years – will yield as well as, if not better than, traditional varieties, and will need 30–50 percent less fertiliser.

But the work is only partly finished for Joko and his Asian partners. They are now building on previous work done at Cornell and EMBRAPA to include the SbMATE gene in their varieties. “Higher yields will only be possible if the plant can also tolerate excess aluminium, which severely inhibits root growth and thereby water and nutrient uptake,” explains Joko. “We are also looking at incorporating salt-tolerance and drought-tolerance genes. It’s a step-by-step process where we hope to build tolerance to the multiple stresses that afflict most rice-growing areas throughout Asia and the world.”

Introducing PSTOL1 into maize and sorghum

At EMBRAPA, Claudia is also interested in building up tolerance to multiple stresses and was involved in the project to look for genes similar to PSTOL1 in maize. “As soon as IRRI and JIRCAS had cloned the gene and created markers, we started using the markers to search for the gene in maize, as Jurandir did in sorghum,” she says.

Women farmers in India bring home their sorghum harvest.

Women farmers in India bring home their sorghum harvest.

Finding genes that confer phosphorus-efficiency traits in maize and sorghum has been a more challenging project, according to Leon. “From the rice work, we knew a big part of phosphorus efficiency was to do with root architecture – you want to have shallow horizontal roots instead of roots that grow down, which is often the case in maize and sorghum,” he explains. “This is because there is less accessible phosphorus further down the soil profile.”

Observing root architecture is difficult in ordinary soil, so the team had to develop new ways to visualise the plants’ roots. They grew plants in a transparent nutrient gel, which they then photographed to create three-dimensional images of the root structure.

The team found sorghum and maize varieties that contained genes similar to PSTOL1 in rice, but which also have longer root systems that radiated outwards rather than downwards in gels with higher concentration of aluminium. “These observations helped validate multiple PSTOL1 regions in sorghum and maize, which we’ve been able to develop markers for to help breeders identify these traits more easily,” says Leon.

These markers have successfully been used by sorghum breeders in Brazil and Africa to identify phosphorus-efficient varieties. Maize breeders in both Brazil and Africa are expected to use similar markers to validate their varieties in 2015.

New sorghum varieties prove their worth in the field

Eva Weltzien is one Africa-based sorghum breeder who has benefited from these PSTOL1 and SbMATE markers. Based in Mali at ICRISAT, Eva and her team have been using the markers to select for aluminium-tolerant and phosphorus-efficient varieties and validating their performance in field trials across 29 environments in three countries in West Africa.

She says the markers have helped evolve the way they do their breeding. “Using molecular markers, we are able to identify whether the lines we are breeding have genes that confer the traits that we want,” explains Eva. “It has really revolutionised our breeding program and helped it make great progress in the past three to four years.”

In Mali, sorghum is an important staple crop. It is used to make (a thick porridge), couscous, and local beers. Part of its popularity is its adaptability to various climates – in Mali it is grown in very dry environments as well as in forest/rainforest zones. However, it is widely affected by acidic soils.

Sorghum farmers at work in the field in Mali.

Sorghum farmers at work in the field in Mali.

“Low phosphorus availability is a key problem for farmers on the coast of West Africa, and breeding phosphorus-efficient crops to cope with these conditions has been a main objective of ICRISAT in West Africa for some time,” says Eva.

“We’ve had good results in terms of field trials. We have at least 20 lines we are field testing at the moment, which we selected from 1,100 lines that we tested under high and low phosphorous conditions.” Eva says that some of these lines could be released as new varieties as early as next year.

“Overall, we feel the GCP partnership with EMBRAPA and Cornell is enhancing our capacity here in Mali, and that we are closer to delivering more robust sorghum varieties that will help farmers and feed the ever-growing population in West Africa.”

Leon notes that the work by Eva in Mali and by other African partners in Niger and Kenya is imperative for the research. “Just because plants have these genes, doesn’t mean they will all display aluminium tolerance or phosphorus efficiency. You still need to test and observe for these traits in the field and determine what other factors might affect plants grown in acidic soils.”

One surprising observation that has Leon intrigued is a local sorghum variety with a phosphorus-efficiency gene that is close to where the SbMATE gene resides in the sorghum genome. “This suggests that SbMATE, which aids with aluminium tolerance, may also improve phosphorus efficiency. This means we could use SbMATE markers to look for both phosphorus efficiency and aluminium tolerance,” he says. Leon and Jurandir will continue to validate this result post-GCP.

Working together to improve food security worldwide

GCP’s comparative genomics projects have laid a significant foundation for further research into and breeding for tolerance to multiple plant stresses.

A Kenyan farmer in her maize field.

A Kenyan farmer in her maize field.

“We’re in a golden age of biology where we are learning more and more about the complexities and commonalities of plants, which is allowing us to manipulate them ever so slightly to help them tolerate multiple environmental stresses,” says Leon. “As a geneticist, I am extremely proud to be part of this, particularly seeing the potential impact that the basic research we do in the laboratory can have on crop improvement and the lives of people in poorer countries.”

Although not all projects produced new and improved varieties ready for release, they are well and truly in the pipeline. Each partner institute is committed to work together and source new funding to continue on their quest to produce further products.

“GCP has really installed in us a spirit to see this work through and expand on it,” says Leon. “I mean, we are now working with other countries and institutes to share what we have learnt with them and help them make the discoveries that we have. It’s a credit to GCP for bringing us all together; that was a key to the success of the project. Each partner has brought their expertise to the table – genomics, molecular biology, plant breeding – and it has been great to see the impact filter into Africa and Asia.”

In Kenya, Samuel agrees with Leon’s assessment. “GCP gave us an opportunity to build our expertise and start interacting with the rest of the world,” he says. “But more importantly, it means that we’re contributing to food security in Kenya, and that makes us really proud.”

Although the sun is setting on GCP, work on comparative genomics projects is still in progress, with all parties still working towards delivering important new acid-beating varieties to farmers.

A boy rides his bicycle next to a rice field in the Philippines. With acid soils affecting half the world’s current arable land, acid-beating crop varieties will help farmers feed their families – and the world – into the future.

A boy rides his bicycle next to a rice field in the Philippines. With acid soils affecting half the world’s current arable land, acid-beating crop varieties will help farmers feed their families – and the world – into the future.

More links