Genetic Maize

The 2010 Maize Genetics Conference started with a call for maize geneticists to take on one of the greatest challenges of human history – feeding the world. Marianne Bänziger of CIMMYT presented the first plenary talk, titled Stress tolerant maize for the developing world – Challenges and prospects. Find the abstract of her talk at the end of this post.

Of all of the staple grains, maize is the most drought susceptible. Wheat is fairly drought tolerant, and rice is irrigated. Maize is sensitive to variation in rainfall, and since it is typically not irrigated, any year to year variation in rainfall will be seen as year to year variation of yield, with low rainfall years yielding less than high rainfall years. There are some drought tolerant varieties that don’t have such variation with rainfall, but they are consistently low yielding, even in high rainfall years. In order to provide enough food for growing populations, maize must be developed that can maintain reasonably high yields even in drought years.

A second major problem with maize is nitrogen. Maize reacts well to fertilizer application, providing (to a point) higher yields with higher amounts of nitrogen. However, less than 50% of applied fertilizer is used by the plant, leaving much of the nitrogen unused This unused fertilizer can be carried via surface waters to places like the Gulf of Mexico where it can contribute to hypoxic zones. Additionally, synthetic nitrogen fertilizer can be expensive to produce because it requires natural gas. Both synthetic nitrogen and non-synthetic fertilizers take fuel to distribute through fields. Maize that can use applied nitrogen more efficiently without laving so much behind must be developed both in order to provide enough food and to ensure that we are using both renewable and nonrenewable resources efficiently while protecting the environment.

CIMMYT aims to solve problems of drought and nitrogen by breeding under stress conditions. Their fields look more like a field in Africa than a field in Iowa. They simply select for stress tolerant plants that grow successfully under low water and low nitrogen conditions. They’ve found that genetic markers in typical yield selected lines and in stress selected lines are very different. CIMMYT is also looking at breeding under low phosphorus and low potassium.

While CIMMYT is primarily focused on breeding, they believe the key to meeting future food needs lies in matching breeding and transgenics. In particular, CIMMYT has partnered with seed companies to develop transgenics that will enhance productivity. The current traits on the market are protective: Bt protects the crop plants from insect damage which can reduce yield, Roundup Ready protects the crop plants from having to compete with weeds for resources, and virus resistance protects papaya from reduced yield due to virus infection. Productivity traits would directly increase yield instead of protecting it. Castigiloni showed in the 2008 paper Bacterial RNA chaperones confer abiotic stress tolerance in plants and improved grain yield in maize under water-limited conditions that yield could be significantly improved with a transgene.

Finding appropriate transgenes that will improve yield isn’t the end of the story. Each transgene needs to be investigated for genotype x environment interactions to see if the productivity transgenes behave differently under different environmental conditions. In addition, the transgenes may behave differently in different varieties, so each individual variety would need to be tested for yield changes with the productivity transgene. More layers of complication are added when multiple genes of similar and different traits are stacked. The combinations of transgenes may behave differently than each gene alone, and the combinations may have different interactions with each variety and environment.

CIMMYT has partnerships with Monsanto to work on water efficient maize for Africa (WEMA) and with Pioneer to work on improved maize for African soils (IMAS) which is also known as nitrogen use efficiency (NUE). These partnerships have many benefits. They can combine CIMMYT germplasm which is adapted for the farming conditions of low-income farms with the elite germplasm held by the corporations. They can ensure that the poor can aces the seed at no cost or at costs they can afford. They can also depend on the companies to provide funding to develop and deregulate the traits.

Developing and using transgenics is not without barriers, of course. In short, transgenics are expensive. Developing a transgenic trait costs $25 to $100 million dollars or more. Costs include finding a gene that does what you want it to, testing efficacy of the gene in many different varieties and environments, safety testing to ensure that the transgenic plants are substantially equivalent to their non-transgenic sister plants, and so on. For the forseeable future, the cost of transgenic traits will remain high. For the price of one commercial transgenic cultivar, CIMMYT believes they can characterize the entire genetic heritage of the two principal cereal crops, wheat and maize.

During her talk, Marianne announced the new CIMMYT program Seeds of Discovery for the first time. This exciting program will examine ancestral varieties of maize and wheat to enable breeding programs globally to use crop biodiversity in developing new lines. They aim to discover the extent of allelic variation in these varieties. They also hope to better understand how the different varieties are related in core sets. Right now, varieties are organized by geographic origin or phenotype but grouping by genotype will allow for better explanation of genetic similarities and differences. When more is known about the allelic diversity in ancestral varieties, marker assisted breeding can be used to bring those rare useful alleles into breeding programs.

In addition to ancestral varieties, CIMMYT looks at farmers’ varieties. They have partners in 14 countries that are both looking for potential lines for breeding that have traits like drought tolerance and looking into how new traits will work with the varieties farmers are currently using. They are also looking into other traits that are important to farmers in the developing world, including taste and appearance.

Greater than 80% of the required yield grain has to come from breeding. No other method, including fertilizer and transgenic traits, will be able to come close to breeding. Making these increases requires scientists from the developed world and from the developing world to both form partnerships and to work on their own areas of expertise. Marianne called upon the maize genetics community to help characterize the genes and alleles that CIMMYT finds in their Seeds of Discovery program. They plan to provide seed to scientists so they can begin to investigate the traits.

Talk Abstract:

Increasing demands for the main food staples, climate change, and increasing water, nutrient and land costs give a new urgency to developing and making available stress tolerant crops. This urgency is the greatest in the developing world where investments in research, capacity building and infrastructure development still lag far behind the developed world. The presentation gives an overview of CIMMYT’s investment in the development of stress tolerant maize which has recently gained significant leverage through stronger research collaboration with public and private partners, and now extends from native and transgenic trait discovery to large scale application of marker assisted selection approaches tailored to the improvement of highly quantitative traits such as yield under drought and low soil fertility. Many years of CIMMYT research indicate that these traits are highly polygenic, which has implications for the use of transgenics, identification of effects within association mapping studies, and the choice of appropriate marker based breeding strategies. In addition to assessing front line transgenics originating from the private sector for use in particular in Africa, current efforts focus on marker assisted recurrent selection (MARS), which is being implemented in over 40 biparental populations in Africa, Asia, and Latin America. Current MARS populations are selected on an index of 200 to 300 anonymous SNP markers, a density chosen because it is affordable with current genotyping technology. In 2010, pilot projects on the implementation of genomic selection (GS) using much higher marker densities will be initiated on new platforms based on next generation sequencing technologies, and it is expected that by 2011 genotyping costs will have dropped enough to permit their routine application across the CIMMYT maize breeding program and facilitate innovative native gene discovery and allele mining approaches. With that, CIMMYT is among the first public sector breeding programs that integrate cutting edge transgenic and molecular techniques on a large scale for germplasm development and dissemination to the tangible benefit of resource poor farmers.

Castiglioni P, Warner D, Bensen RJ, Anstrom DC, Harrison J, Stoecker M, Abad M, Kumar G, Salvador S, D’Ordine R, Navarro S, Back S, Fernandes M, Targolli J, Dasgupta S, Bonin C, Luethy MH, & Heard JE (2008). Bacterial RNA chaperones confer abiotic stress tolerance in plants and improved grain yield in maize under water-limited conditions. Plant physiology, 147 (2), 446-55 PMID: 18524876

Volker Brendel, professor of bioinformatics at Iowa State, spoke at the Maize Genetics Conference about the need for a better system of community annotation of the maize genome. The genome of the popular maize inbred line B73 is sequenced, but we don’t actually know what a lot of the code stands for. It’s going to take a lot of collaborative effort to discover and annotate (explain) the function of each gene and to put all of that information in one place so it will be useful.
Volker reminds us that the Arabidopsis 2010 funding is running out, so we need to assess the plant genetics situation. How many genes do we know the function of? There is still much to learn.
Maize is uniquely positioned to replace Arabidopis as a focus for basic plant research due to the many resources that are already established, the most important of which is the extensive maize genetics community (he didn’t say it, but there is another reason why maize is a better choice than Arabidopsis right now – all of our major grains are very closely related, so work on maize applies to rice, wheat, sorghum, and more). The community needs to work together in the annotation process, assigning functions to the genes that have been sequenced, putting the data from a variety of sources together to make a bigger picture. Each researchers has a favorite gene (pathway, organelle, etc) – how can each of the researchers contribute to the annotation process?
PlantGDB is a comparative genomics site funded by NSF has information on 14 species, including maize, which is very useful. However, no matter how clever the computer programs are, the human touch is still needed. Filling in information on any of these species helps us to better understand all of them. On the site, community members can flag genes for which the models don’t seem to fit, and can contribute alternative explanations. The final goal is to have every gene model approved by the relevant community member(s). When a person annotates a gene, the PlantGDB committee reviews it, approves it, and the information is shortly available on the site. Annotating the genes you are working on is your civil duty, something you owe due to public funding you receive.
After Volker’s talk, the attendees discussed what is the public’s role in the attenuation process should be. There are a lot of cases where the the gene model can be checked without any lab work, simply by looking at the sequences. Some members of the community think we should harness the brainpower of thousands of biology undergraduate students by assigning annotations for class. I like the idea of getting students involved, and hope they follow through.Diversity of people to represent the maize genetics community.
A panel discussion followed, where a lot of great new ideas for annotation were brought up (unfortunately I don’t have the names of some of the people that spoke).
One panel member said we need “Zeazomics” – a collection of information including genomics, metabolimics, proteomics, and whatever else we can come up with – to fill in gaps in our knowledge. being able to link all of this information together will lead to stronger explanations of the phenotypes we see. He said this process will not be definitive, it will create a series of hypothesis that will lead to more hypotheses. The hypothesis testing will lead to functional biolgoy, from physiology to biochemistry to cell biology and more. Additional genome sequencing is necessary to capture the entire diversity of maize. Maize is the model for grasses, for crops, for future applications like biofuels. Now is the time to push maize research to a much higher level.
To accomplish all this, we’ll need to take care of a few things, as the other panel members and members of the community brought up:

• Need to have reciprocal links from genes from MaizeGDB to NCBI Entrez Gene. Currently, about 20,000 NCBI Entrez Genes need links back to MaizeGDB.
• To help with annotation, Lisa Harper, curator of MaizeGDB, will do a movie that shows the common problems of using the databases, including how the genome changes over time as the contigs are reordered, etc. This is needed because people are often working off of older copies of the information for a given gene, as it might not be updated frequently enough.
• There is also a need to integrate microarray data into the databases. Particularly complicated are those microarrays that are specific to a particular tissue and/or developmental stage. Volker says that this problem is common and new technologies with new ways to visualize data are necessary.
• MaizeGDB needs a forum such that people working on the same genes can coordinate their work.
• iPlant is organizing a workshop in St. Louis in June to help coordinate the various genome annotation groups.

• There is a plan to create outreach information that any member of the maize community will be able to download and use to communicate the needs and accomplishments to the public and to government officials.

The 51st Maize Genetics Conference is just as overwhelming as I remembered from the the 49th (50 was in Washington DC and was too expensive for me to go). We have 480+ maize geneticists all in one resort in St. Charles, IL, presenting 244 posters, 4 plenary talks, 35 short talks, and innumerable conversations about maize. The topics range from perfecting the corn genome sequence to writing and using software to help us navigate it to the intricate details of transposons and centromeres… and a little bit of applied work as well. There’s no way one person could convey all of the information presented here, but I hope I can share some of the tidbits that were particularly interesting to me in my next few posts. You can find the entire 201 page program on the MaizeGDB website, which includes abstracts of all of the posters and talks. Many people will upload their talks and posters to the website, I’ll let you know when they go up.

The NCCC-167 meeting is over, and I’m very glad to have had the opportunity to attend. The acronym stands for North Central Communications Committee, and 167 is the USDA-ARS project number. It turns out that there are hundreds of projects, some of which are designated for conferences and communications, such as this one. Apparently this particular conference used to be NCR-2 (North Central Region) but the rumor is that a Kansas corn breeder forgot to renew the project in time, so the group had to reapply and got a much higher number. That happened so long ago that 167 is a well recognized number in the corn breeding community.
The most important idea I took from the meeting, besides the reminder that there’s a lot more to scientists than you’d think from just reading their papers (as I described in NCCC-167), is that groups really need to stay organized. The Maize Genetics Community is well structured and very large, as can be seen from the huge number of attendees at the Maize Genetics Conference (more on that in another post, it’s where I am at this very moment). There are just as many if not more maize breeders as there are maize geneticists, but they don’t have as cohesive of a community. I’m not sure why this is, but it certainly seems to be a problem. Without strong lines of communication across the community, the group has a decreased ability to apply for collective grants, less ability to share information and techniques, etc. The importance of breeding will only grow as climate change brings diseases and pests to areas where they did not exist before and as population growth demands higher yields. My major professor and  a few others in the community seem dedicated to bringing stability and continuity to the group. I look forward to watching it grow back to its former glory.
I took extensive notes on the wonderful talks at NCCC-167, and hope to post more about them in the coming days. Most importantly, I’m anticipating additional information about breeding high methionine maize for organic chicken feed from Walter Goldstein of the Michael Fields Agricultural Institute. We may not see eye to eye on every topic, but I certainly agree that we could use nutritionally enhanced corn, and that’s one heck of an important place to start a conversation!
For now, I must turn my attention to the Maize Genetics Conference – let’s hope I can keep up!

Shawn Kaeppler is a researcher at the Great Lakes Bioenergy Center, a DOE Bioenergy Research Center, and the University of Wisconsin Department of Agronomy. Specific to this center is a lot of work on sustainability, including work on energy balance. You may be surprised, then, to hear that Shawn’s group is working on corn stover, but they have a strong rationale for doing so. Corn is closely related to the potential biofuel perennials miscanthus and switchgrass. They work in corn because of the resources available like a sequenced genome and large germplasm sources, and will then use the knowledge they acquire to improve the grasses.
Groups within the Great Lakes Bioenergy Center and the Agronomy Department of the University of Wisconsin are working on determining how much stover can be removed from the field. Some must remain to prevent erosion and to contribute to soil carbon. There is no easy answer, as the amount you can remove depends on soil quality, slope of the field, farming practices, and more. Some areas will need to retain more stover than others.
Core to biofuel research (as with most other breeding efforts) is screening diverse germplasm for candidate genes that correlate with traits like biomass yield and ethanol potential. Shawn’s group uses three main methods: microarrays in specific tissues, QTL analysis, and searching for candidate genes directly.
Specific traits under analysis fall under biomass quality and biomass quantity. Quality traits include using ruminant digestibility parameters because these qualities seem to be quite related to biofuel production, and provide an overall picture rather than focusing on one biochemical pathway. Quantity traits include stalk diameter and internode length.
One trait that might be related to quality is vegetative phase change, which refers to the time when juvenile tissues transition to adult tissues. Phase change timing is a trait susceptible to selection. This research addresses how timing differentials are inherited and whether having more juvenile tissue positively effects biomass quantity and quality. They found no relationship between the time of phase change and digestibility of the whole plant (without grain), however there is a relationship (although not significant) between the proportion of leaves that are still juvenile and digestibility. Overall, it seems that increasing juvenile tissue does not increase digestibility and also reduces yield because the tissue is more susceptible to disease and insects, and may senesce before harvest. This is good to know, because researchers can move on to traits that do affect digestibility / biomass quality.
Candice Hansey is a student of Shawn Kaeppler at the University of Wisconsin studying biomass quality. Her work answers the question: how can we measure ethanol potential? Simultaneous saccharification and fermentation is a great method, but very low throughput (takes a whole day to do 15 samples). Instead, they use established methods to measure forage quality (digestibility). The samples are put into a packet of filter paper, weighed, then subjected to a succession of different solvents that dissolve different fractions of the samples which then leave the filter packet. For example, the last solvent is 72% sulfuric acid which dissolves everything except acid-insoluable lignin. With this method, they can determine what percentage of each sample is lignin, carbohydrate, etc. Candy has screened different plant parts at different levels of maturity to see which might best predict the digestibility of the total adult plant. Unfortunately, the most predictive tissue was the stalk at developmental phase R6, but dissecting adult plants is a hassle. She determined that whole plant analysis is the best way to screen for digestibility.
These projects are great example of how negative results can be just as useful in moving science forward as positive results. Thanks to Shawn and Candy, we now know some areas of research that can be “checked off”, so to speak, allowing resources to be used in other areas.
Note: this post consists of my notes of Shawn and Candy’s talks at NCCC-167 2009, and was posted with their agreement.