More Sustainable Feedstock for Ethanol

A six-year Great Lakes Bioenergy Research Center (GLBRC) study on the viability of different bioenergy feedstocks recently demonstrated that perennial cropping systems such as switchgrass, giant miscanthus, poplar, native grasses and prairie can yield as much biomass as corn stover.

The study is significant for addressing one of the biofuel industry’s biggest questions: Can environmentally beneficial crops produce enough biomass to make their conversion to ethanol efficient and economical?

Since 2008, research scientists Gregg Sanford and Gary Oates, based in the lab of CALS agronomy professor Randy Jackson, have worked with colleagues at Michigan State University (MSU) to cultivate more than 80 acres of crops with the potential to become feedstocks for so-called “second-generation” biofuels, that is, biofuels derived from non-food crops or the nonfood portion of plants. They’ve grown these crops at the CALS-based Arlington Agricultural Research Station and at MSU’s Kellogg Biological Station.

“We understand annual systems really well, but little research has been done on the yield of perennial cropping systems as they get established and begin to produce, or after farmland has been converted to a perennial system,” says Oates.

To find out basic information about how well certain crops produce biomass, Sanford and Oates tested the crops across two criteria: diversity of species, and whether a crop grows perennially (continuously, year after year) or annually (needing to be replanted each year).

Highly productive corn stover has thus far been the main feedstock for second-generation biofuels. And yet perennial cropping systems, which are better equipped to build soil quality, reduce runoff, and minimize greenhouse gas release into the atmosphere, confer more environmental benefits.

Corn, when grain is included, proved to be most productive over the first six-year period of the study at the Wisconsin site, but giant miscanthus, switchgrass, poplar and native grasses were not far behind. At the MSU site, where soil is less fertile, miscanthus actually produced the same amount of biomass as corn (grain included) in the experiment, with poplar and switchgrass within range.

“All of this means that, at large scales and on various soils, these crops are competitive with corn, the current dominant feedstock for ethanol,” Sanford says.

Now in the midst of the study’s eighth year, Sanford says the study will continue for the foreseeable future.

“We know that perennial systems can prevent negative impacts such as soil erosion and nitrate leaching, and that they also provide habitat for native species that provide beneficial ecosystem services,” Sanford says. “But there are still a lot of questions we want to answer about soil processes and properties— questions that take many years to answer.”

Researcher Gregg Sanford stands before a plot of giant miscanthus at Arlington.

Photo credit – Matthew Wisniewski

The Future, Unzipped

John Ralph PhD’82 talks with the easy, garrulous rhythms of his native New Zealand, and often seems amiably close to the edge of laughter.

So he was inclined toward amusement last year when he discovered that some portion of the Internet had misunderstood his latest research. Ralph—a CALS biochemist with joint appointments in biochemistry and biological systems engineering—had just unveiled a way to tweak the lignin that helps give plants their backbone. A kind of a natural plastic or binder, lignin gets in the way of some industrial processes, and Ralph’s team had cracked a complicated puzzle of genetics and chemistry to address the problem. They call it zip-lignin, because the modified lignin comes apart—roughly—like a zipper.

One writer at an influential publication called it “self-destructing” lignin. Not a bad turn of phrase—but not exactly accurate, either. For a geeky science story the news spread far, and by the time it had spread across the Internet, a random blogger could be found complaining about the dangers of walking through forests full of detonating trees.

Turning the misunderstanding into a teachable moment, Ralph went image surfing, and his standard KeyNote talk now contains a picture of a man puzzling over the shattered remains of a tree. “Oh noooo!” the caption reads. “I’ll be peacefully walking in a national park and these dang GM trees are going to be exploding all around me!”

That’s obviously a crazy scenario. But if the technology works as Ralph predicts, the potential changes to biofuels and paper production could rewrite the economics of these industries, and in the process lead to an entirely new natural chemical sector.

“When we talk to people in the biofuels industry, what we are hearing is that creating value from lignin could be game-changing,” says Timothy Donohue, a CALS professor of bacteriology and director of the UW–Madison-based Great Lakes Bioenergy Research Center, where Ralph has a lab. “It could be catalytic.”

After cellulose, lignin is the most abundant organic compound on the planet. Lignin surrounds and shapes our entire lives. Most of us have no idea—yet we are the constant beneficiaries of its strength and binding power.

When plants are growing, it’s the stiffening of the cell wall that creates their visible architecture. Carbohydrate polymers—primarily cellulose and hemicelluloses—and a small amount of protein make up a sort of scaffolding for the construction of plant cell walls. And lignin is the glue, surrounding and encasing this fibrous matrix with a durable and water-resistant polymer—almost like plastic. Some liken lignin to the resin in fiberglass.

Without lignin, the pine cannot soar into the sky, and the woody herb soon succumbs to rot. Found primarily in land plants, a form of lignin has been identified in seaweed, suggesting deep evolutionary origins as much as a billion years ago.

“Lignin is a funny thing,” says Ralph, who was first introduced to lignin chemistry as a young student during a holiday internship at New Zealand’s Forest Research Institute. “People who get into it for a little bit end up staying there the rest of their lives.”

The fascination is born, in part, from its unique chemistry. Enzymes, proteins that catalyze reactions, orchestrate the assembly of complex cell wall carbohydrates from building blocks like xylose and glucose. The types of enzymes present in cells therefore determine the composition of the wall.

Lignin is more enigmatic, says Ralph. Although its parts (called monomers) are assembled using enzymes, the polymerization of these parts into lignin does not require enzymes but instead relies on just the chemistry of the monomers and their radical coupling reactions. “It’s combinatorial, and so you make a polymer in which no two molecules are the same, perhaps anywhere in the whole plant,” says Ralph.

This flexible construction is at the heart of lignin’s toughness, but it’s also a major obstacle for the production of paper and biofuels. Both industries need the high-value carbohydrates, especially the cellulose fraction. And both have to peel away the lignin to get to the treasure inside. A combination of heat, pressure, and caustic soda is standard procedure for liberating cellulose to make paper; bleach removes the remaining lignin. In the biofuels industry, a heat and acid or alkaline treatment is often used to crack the lignin so that it is easier to produce the required simple sugars from cellulose. Leftover lignin is typically burned.

The economic cost of these treatments alone is significant, and lignin pretreatment is at the heart of many of the more egregious environmental costs of paper. On the biofuels side, lowering treatment costs to liberate carbohydrates from lignin could change the very economics of biofuels. In these large-scale, industrial processes, saving a percentage point or two is often worthwhile, but the Holy Grail is a quantum jump.

“Because it’s made this way”—Ralph jams his hands together, crazy-wise, fingers twisted together into a dramatic representation of lignin polymerization—“there is no chemistry or biology that takes it apart in an exquisite way,” he says. “We actually stepped back and thought: How would we like to design lignin? If we could introduce easily cleavable bonds into the backbone, we could break it like a hot knife through butter. How much can you actually mess with this chemistry before the tree falls down?”

Ralph’s team had their eureka moment more than 15 years ago, and have been trying to bring it to life ever since.

With a background in forage production and ruminant nutrition, John Grabber, an agronomist at the USDA–Agricultural Research Services’ Dairy Forage Research Center in Madison, got pulled into lignin chemistry through the barn door. On his family’s dairy farm he grew up with lignin stuck to his boots, though he never knew it. But during graduate school he became interested in how plants are digested by cows. Cell walls are potentially a great source of digestible carbohydrates—most plants contain anywhere from 30 to 90 percent of their mass in their cell walls—but it is entangled with lignin. “You quickly find out that lignin is the main barrier to feed digestion,” he says.

Grabber began working on a model system to understand plant lignification—for corn in particular—in 1989. After meeting at a conference, Grabber joined Ralph and plant physiologist Ronald Hatfield at the Dairy Forage Research Center back in 1992. There were many projects ongoing, but Grabber remained interested in trying to fully understand the structural characteristics of the lignin: how it’s made and how to modify it. In his model system they could make any kind of lignin they wanted to study, and see how the changes affected utilization.

Ralph and Hatfield advocated for the work, helping to find funding and offering their expertise. “If I had worked for somebody else I probably wouldn’t be doing this work,” says Grabber. “John and Ron gave me freedom and support to do it.”

Around the same time, Fachuang Lu joined Ralph’s lab seeking a Ph.D. His journey into lignin chemistry was not, at first, his idea. A native of mainland China, he’d enjoyed a successful undergraduate career in Beijing studying chemical engineering, then found himself assigned by the college to a master’s program in lignin chemistry. Lignin is an ingredient in the slurry of chemicals used in oil drilling, and that was his specialty. In 1989 Lu left Beijing for a teaching position at Guangxi University, but three years later he decided to continue his education. Though he’d never met Ralph, he was fascinated by the chemistry and applied to study in his lab.

As Ralph, Grabber, Hatfield and Lu continued to tinker with lignin chemistry, momentum began to build in the lab. Though lignin created a snowflake universe of different molecules, there were rules of assembly. A complex chemical pathway enabled lignin construction, with a mechanism that remained constant across different families of plants, but with many potential building blocks.

Ralph and his colleagues were the first to detail what was happening to lignin as the controlling genes of the biosynthetic pathway were turned on and off, a task ably completed by a slew of outstanding collaborators worldwide with expertise in biotechnological methods—but who lacked the diagnostic structural tools to determine what the plant was doing in response.

Ralph’s team quickly learned that lignification was somewhat flexible. “We figured that we could engineer lignin well beyond the previously held bounds,” says Ralph. As various pathways and chemical possibilities danced in their heads, it struck them: What if, during lignification, they could persuade the plant to slip in a few monomers that had easily broken chemical bonds? If they did it right, lignin would retain its structural value to the plant, but be easier to deal with chemically.

“In the course of our conversation we realized that if plants could do this, it could really revolutionize how readily you could make paper,” recalls Grabber. Says Ralph: “It’s almost impossible to tell which one of us actually verbalized it first—it is one of those great outcomes of the group dynamic.”

Lu’s particular genius was synthesizing the various complex chemicals needed, particularly a novel monomer-conjugate called coniferyl ferulate. It was the key to the zip-lignin—the teeth of the zipper. “He’s got to be one of the best in terms of making molecules,” says Grabber.

They were thrilled by such a revelation, but, in retrospect, they soon realized it was sort of an obvious idea—one suggested by the underlying chemistry and biochemistry of a pathway that was becoming increasingly well understood. Yet it was a discovery of huge potential value. They dropped into stealth mode and began to work on it. They finished important research and stuck it in drawers—signature research, the kind that, when finally published, would capture journal covers. And yet they sat on it, quietly chipping away for nearly a decade.

It helped that there was a flurry of controversy in the field—what Chemical & Engineering News called “the lignin war.” “Part of the reason we could sit on it was that, at the time, making these kinds of molecules was so far-fetched,” says Grabber. “Probably if we had talked about it, people would have laughed at us.”

But as the idea for zip-lignin grew in principle, it became stronger. Lu, Hatfield and colleague Jane Marita MS’97 PhD’01 found that balsa trees and a fiber crop known as kenaf produced very small amounts of coniferyl ferulate. But even as the idea seemed more and more feasible, Hatfield and Marita couldn’t isolate the gene needed to manufacture coniferyl ferulate because of its very low expression in these plants.

And they got stuck. “At the beginning we were thinking that this is just a fantastic idea, but we really didn’t have that much confidence,” says Lu. “Maybe John [Ralph] had more confidence than me.” So they just kept at it. “Every step you think, yes, we are closer, closer, closer.”

In 2008 Ralph moved his work from the Dairy Forage Research Center into UW labs, with research projects under the recently formed Great Lakes Bioenergy Research Center (GLBRC). The center, launched with a $125 million grant from the U.S. Department of Energy that has since been renewed, was just one manifestation of the money and intellectual heft infusing biofuels research—and for zip-lignin it was a lucky move.

During the center’s first full meeting, Curtis Wilkerson, a plant biologist at GLBRC partner Michigan State University, was sitting in the audience when Ralph took his turn at the podium.

Wilkerson is a cell wall specialist. Though lignin is a third of the wall’s carbon and is essential to the way plants conduct water, he confesses he’d never given it much thought. In a room full of cell wall specialists, Ralph would “likely be the only person talking about lignin,” he says. “It just split that way a long time ago. People like myself had very little exposure to what John was thinking.”

It was this kind of academic silo that a place like GLBRC was supposed to breach. Ralph talked about putting ester bonds into lignins and his team’s long search for the elusive enzyme. Wilkerson saw a solution. Due to recent technical advances, the price of determining all of the expressed enzymes in a plant had become more refined and much less expensive. He offered to use these recent developments to try to find the missing enzyme to enable zip-lignin.

From the previous work, Wilkerson knew essentially the size and shape of the puzzle piece he was looking for. He began, quite literally with Google, trolling through the scientific literature looking for a plant that made a lot of coniferyl ferulate. The Chinese medicinal “dong quai” or Chinese angelica (Angelica sinensis) soon emerged as a candidate. Its roots contained about 2 percent coniferyl ferulate.

The team used knowledge about the likely type of enzyme they were searching for and successfully identified the gene and its enzyme that could produce coniferyl ferulate. The whole search took less than six months.

Would you believe an essential tool for the genetic engineering of poplars is a hole punch? That’s the word from Shawn Mansfield, a molecular biochemist at the University of British Columbia, who took the zip-gene from the Angelica and made it work in poplar, a popular tree in the biomass and forest products industry.

Working from Wilkerson’s gene, the first job was figuring out how to tag the new protein so that it fluoresced during imaging. While not necessary to the function of the genetically modified plant, it essentially allows the scientists to check their work: see where the protein is, how much is there, and if it is behaving as a protein should.
Mansfield’s lab also had to find a way to turn the gene on at the right time and place. It could make all the coniferyl ferulate one wanted, but if it wasn’t made at the right time and tissue, there would be no zip-lignin.

After perfecting these finer points, the gene is inserted into a special bacterium—and then the hole punch finally comes into play. Disks punched from poplar leaves are mixed with bacteria that have been inoculated with a special chemical that stimulates the bacteria to share their DNA around. Then the leaf disks are put in a special growth medium. As many as 12 shoots might emerge off of a single disk, but the lab would select and nurture only one shoot from each disk.

In the end they had about 15 successful transgenic candidates that they grew in the greenhouse and then shipped off to Wilkerson and Ralph for further study. Final selection was made based on the amount of fluorescent yellow the trees gave off, and from a newly devised analytical method developed by Lu and Ralph that was particularly diagnostic for the incorporation of the zip monomer into the lignin polymer.

The team knew that genetically modified organisms are not popular or easily talked about—never mind the exploding trees. The idea of reworking a fundamental building block of the plant world will breed resistance.

Ralph argues that this is already part of nature’s vocabulary: they’ve found their building blocks within the plant kingdom, including mutants that do similar things. And now that they know what they are looking for, Steven Karlen, a member of Ralph’s group, is continuing to find more evidence that Mother Nature is doing it herself. “We managed to persuade plants to do this,” Ralph says. “Chances are that nature has already attempted it and you could actually get there by breeding.”

It’s no surprise that Mansfield, who created the final transgenic tree, argues that there is a role for this kind of technology. “We as scientists should be wise in advocating for the proper use of it,” he cautions. “I would never force it on anybody. I would never try to sway people to think that it is the end-all or be-all for everything.”
But given the growing human population and rising CO2 levels, something like zip-lignin has a definite use in reducing the carbon footprint by reducing processing energy and chemical loads. “That means there are less environmental pollutants that need to be cleaned up afterwards,” Mansfield says.

“Our ecological footprint can be much reduced using these kinds of transgenic trees,” he argues. “The caveat is that we need to be very smart about where and how we plant them.”

Not many things in the natural world can take apart lignin, but any homeowner with a deck knows that fungi are up to the task. A recent analysis of mushroom genomes suggests that fungi evolved this ability about 300 million years ago. This is about the end of the Carboniferous era, when earth’s coal production began to slow down. Coincidence? Perhaps not. Now that wood could rot, it probably slowed the burial of organic carbon via tree trunks and other lignin-rich plants.

Could the discovery of zip-lignin signal another transition, and hasten our move away from fossil fuels laid down in the Carboniferous?

Tim Donohue likes to think so. He likens biofuels now to the early oil industry, when oil was simply being turned into liquid fuel while the by-products were burned or dumped. It took a few decades for inventors to capitalize on this now valuable stream of raw materials to build the modern chemical industry.

“Lignin is about 25 to 30 percent of carbon in the plant. So if we’re going to catalyze an industry that makes clean energy and chemicals from plant biomass, figuring out what to do with the lignin is going to be key,” Donohue says.

People in the industry used to joke that you could do a lot of things with lignin except make money from it. But that may be changing. “The economics and profitability of the industry will be very different if lignin can be turned into valuable compounds,” says Donohue.

One of the early efforts to make use of lignin was in Rothschild, Wisconsin, at a company now known as Borregaard LignoTech. When processed properly, lignin has many uses, from the manufacture of vanilla flavor to additives for concrete. There is even a small amount of it in the battery of your car that allows it to keep recharging.

Jerry Gargulak is research manager at Borregaard LignoTech, and learned about zip-lignin recently in his capacity as a scientific advisor to the GLBRC. Despite its many uses, Gargulak and his colleagues dream about a time when lignin can replace carbon black in tires and be used to build carbon fibers and structural plastics.

Zip-lignin and the ideas behind it could bring this day closer. “It gives us a technology that might yield a more interesting lignin-derived starting material,” Gargulak says. “It could potentially lead to a lot of innovation downstream in lignin technology.” But he emphasizes, “There are a lot of i’s to be dotted and t’s to be crossed.”

This story is just beginning. Zip-lignin has a patent and has excited industrial interest that could be worth significant dollars. Ralph and his colleagues continue working to further refine the process, increasing the percentage of zippable bonds in poplar and also inserting the gene into more plants, such as corn and Brachypodium, both grasses.
And in the basement of the shiny new Wisconsin Energy Institute building, where the GLBRC is based, two massive new nuclear magnetic resonance (NMR) spectrometers work 24/7, providing a level of detail into lignin that Ralph has never had before.

“We spend a lot of time looking at these Rorschach test–like figures,” Ralph says of the information generated from the NMR. “The detail in them is unbelievable. These things have been revolutionizing what we do.”

Many Paths of Discovery

Having an applied research goal can no doubt lend focus to the discovery process. For example, since its inception the charge of the Great Lakes Bioenergy Research Center here on campus, funded by the U.S. Department of Energy, has been to realize the grand vision of a biorefinery—the bioenergy version of the petroleum refinery. If we’re investigating biomass as a source of material that we’re going to get products from, we need to understand both how it’s put together and how to take it apart.

This quest has generated discoveries great and small, including CALS biochemistry professor John Ralph’s groundbreaking work in technologies to take apart lignin, a particularly tough compound in plant cell walls.

But pioneering discoveries don’t always happen with a specific application in mind—or applications are later found that are bigger and bolder than the researcher could originally conceive of. Take, for example, the late CALS
genetics professor Ray Owen’s investigation of twin calves with different fathers that somehow were able to tolerate carrying each other’s differing blood cells—a mix that often triggers a severe immunological reaction. But when blood cells are exchanged early in development, Owen learned, each twin learns to tolerate the other’s cells.

By asking questions about a common occurrence in cattle, Owen had discovered the phenomenon of immune tolerance, which sparked a revolution in immunology and laid the foundation for the successful transplantation of human organs. His findings, published in 1945, paved the way for research involving induction of immune tolerance to transplanted tissue grafts by Frank Burnet and Peter Medawar. When those scientists received the Nobel Prize for that work in 1960, they noted it was Owen’s discovery that had set them on their way.

For another example, fast-forward to the present and consider the research of plant pathologist Aurélie Rakotondrafara, highlighted in our Grow cover story. While pursuing a basic science question—how plant viruses reproduce—she happened upon a very useful tool: a stretch of genetic material in a plant virus, known as an “IRES,” that is powerful at “recruiting” the plant’s natural machinery for making proteins.

It turns out there are huge biotech applications for this finding. “Rakotondrafara wasn’t looking for a more efficient tool to make proteins, but the IRES she found is perfect for it,” notes Jennifer Gottwald, a technology officer at the Wisconsin Alumni Research Foundation, which is working on a patent for this discovery.

That’s the excitement of scientific curiosity—and the best reason why we place such high value on both basic and applied research. One feeds into the other, and we cannot fully know the potential outcomes of discoveries we make today. We actively foster this curiosity about how living things work because the fruits of research are boundless, and often yield tremendous unexpected gifts along the way.

Uganda: The Benefits of Biogas

Generating enthusiasm for a new kind of technology is key to its long-term success. Rebecca Larson, a CALS professor of biological systems engineering, has already accomplished that goal in Uganda, where students at an elementary school in Lweeza excitedly yell “Biogas! Biogas!” after learning about anaerobic digester systems.

Larson, a UW–Extension biowaste specialist and an expert in agricultural manure management, designs, installs and upgrades small-scale anaerobic digester (AD) systems in developing countries. Her projects are funded by the Wisconsin Energy Institute at UW–Madison and several other sources. Community education and outreach at schools and other installation sites are an important part of these efforts.

Children get excited by the “magic” in her work, she says. “It’s converting something with such a negative connotation as manure into something positive,” Larson notes. In an AD system, this magic is performed by bacteria that break down manure and other organic waste in the absence of oxygen.

The resulting biogas, a form of energy composed of methane and carbon dioxide, can be used directly for cooking, lighting, or heating a building, or it can fuel an engine generator to produce electricity.

Larson’s collaborators in Uganda include Sarah Stefanos and Aleia McCord, graduate students at the Nelson Institute for Environmental Studies who joined forces with fellow students at Makarere University in Kampala to start a company called Waste 2 Energy Ltd.
Along with another company, Green Heat Uganda, which has built a total of 42 digesters, Waste 2 Energy has helped install four AD systems since 2011.

“Most of these digesters are locally built underground dome systems at schools and orphanages,” Larson explains. Lweeza’s elementary school is a perfect example.

The AD systems use food waste, human waste from pit latrines and everything in between. The biogas generated by the digester is run through a pipeline to a kitchen stove where the children’s meals are prepared. Compared to traditional charcoal cooking, the AD systems greatly reduce the school’s greenhouse gas emissions.

Larson and her team are now focusing on enhancing the efficiency and environmental benefits of these systems. Their goals are to improve the digester’s management of human waste, reduce its water needs, increase the amount of energy it produces and generate cheap fertilizer to boost food crop yields.

“Our overall goal is to create a closed-loop and low-cost sustainability package that addresses multiple local user needs,” Larson says.

The beauty of the project is that all these needs can be met by simply adding two new components to the existing systems: heating elements and a solid-liquid separator.

To help visualize the impact of the fertilizer, Larson set up demonstration plots that compare crop yields with and without it. Down the road, a generator could be added to the system to provide electricity in a country where only 9 percent of the population currently has access.

As a next step, Larson hopes to replicate the project’s success in Bolivia. She is finalizing local design plans with Horacio Aguirre-Villegas, her postdoctoral fellow in biological systems engineering, and their collaborators at the Universidad Amazonica de Pando in Cobija.

Forever Rising

To begin to understand the outsized potential and sheer weirdness of yeast, it helps to consider the genetics behind one of the world’s most successful and useful microorganisms. It also helps to consider lager.

Lager, or cold-brewed beer, is made possible by the union of two distinct species of yeast. About 500 years ago, these two species, Saccharomyces eubayanus and Saccharomyces cerevisiae, joined in a Bavarian cellar. They gave us a hybrid organism that today underpins an annual global market for lager estimated at one-quarter of a trillion dollars.

“We would not have lager if there hadn’t been a union equivalent to the marriage of humans and chickens,” notes Chris Todd Hittinger PhD’07, a CALS professor of genetics and a co-discoverer of S. eubayanus, the long-sought wild species of yeast that combined with the bread- and wine-making S. cerevisiae to form the beer. “That’s just one product brewed by one interspecies hybrid.”

Yeasts, of course, are central to many things that people depend on, and the widespread domestication in antiquity of S. cerevisiae is considered pivotal to the development of human societies. Bread and wine, in addition to beer, are the obvious fruits of taming the onecelled fungi that give us life’s basics. But various strains and species of yeasts also are partly responsible for cheese, yogurt, sausage, sauerkraut, kimchi, whiskey, cider, sake, soy sauce and a host of other fermented foods and beverages.

Baker’s yeast, according to yeast biologist Michael Culbertson, an emeritus professor and former chair of UW– Madison’s Laboratory of Genetics, ranks as “one of the most important organisms in human history. Leavened bread came from yeast 5,000 years ago.”

Beyond the table, the microbes and their power to ferment have wide-ranging applications, including in agriculture for biocontrol and remediation, as well as for animal feed and fodder. They are also widely used to make industrial biochemicals such as enzymes, flavors and pigments.

What’s more, yeasts are used to degrade chemical pollutants and are employed in various stages of drug discovery and production. Human insulin, for instance, is made with yeast. By inserting the human gene responsible for producing insulin into yeast, the human variant of the hormone is pumped out in quantity, supplanting the less effective bovine form of insulin used previously.

Transforming corn and other feedstocks, such as woody plant matter and agricultural waste, to the biofuel ethanol requires yeast. Hittinger is exploring the application of yeast to that problem through the prism of the Great Lakes Bioenergy Research Center (GLBRC), a Department of Energy-funded partnership between UW–Madison and Michigan State University. Hittinger leads a GLBRC “Yeast BiodesignTeam,” which is probing biofuel applications for interspecies hybrids as well as genome engineering approaches to refine biofuel production using yeasts.

“There are lots and lots of different kinds of yeasts,” explains Hittinger. “Yeasts and fungi have been around since Precambrian time—hundreds of millions of years, for certain. We encounter them every day. They’re all around us and even inside us. They inhabit every continent, including Antarctica. Yeasts fill scores of ecological niches.”

The wild lager beer parent, S. eubayanus, for example, was found after a worldwide search in the sugarrich environment of Patagonian beech trees—or, more specifically, in growths, called “galls,” bulging from them. (How S. eubayanus got to Bavaria hundreds of years ago and made the lager hybrid possible remains a mystery.) It is possible, notes Hittinger, to actually smell the S. eubayanus yeast at work, churning alcohol from the sugars in the galls themselves.

Though the merits of known yeast species for making food, medicines and useful biochemicals are numerous, there are likely many more valuable applications of existing and yet-to-bediscovered yeasts.

For Hittinger and the community of yeast biologists at UW–Madison and beyond, a critical use is in basic scientific discovery. The use of yeast as a research organism was pioneered by Louis Pasteur himself, and much of what we know about biochemical metabolism was first studied in yeasts.

Since the 1970s, the simple baker’s variety of yeast has served as a staple of biology. Because yeasts, like humans and other animals, are eukaryotes— organisms composed of cells with a complex inner architecture, including a nucleus—and because of the ease, speed and precision with which they can be studied and manipulated in the lab, they have contributed significantly to our understanding of the fundamentals of life. And because nature is parsimonious, conserving across organisms and time useful traits encoded as genes, the discoveries made using yeast can often be extended to higher animals, including humans.

“The model yeast, S. cerevisiae, has been instrumental in basic biology,” says Hittinger. “It has told us something aging. In terms of understanding basic processes, it’s a tough model system to beat. It’s a champion model organism for genetics and biochemistry.”

“It is widely unappreciated how thevast terrain of biology has been nourished by yeast,” argues Sean B. Carroll, a CALS professor of genetics and one of the world’s leading evolutionary thinkers. It was in Carroll’s lab a decade ago as a graduate student that Hittinger first turned his attention to yeast, coauthoring a series of high-profile papers that, among other things, used the yeast model to catch nature in the act of natural selection, the proof in the pudding of evolutionary science.

Now the model is about to shift into an even higher gear. The work of Hittinger and others is poised toenhance the yeast model, add many new species to the research mix, and begin to make sense of the evolutionary history of a spectacularly successful and ubiquitous organism. The advent of cheap and fast genomics—the ability to sequence and read the DNA base pairs that make up the genes and genomes of yeasts and all other living organisms—along with the tools of molecular biology and bioinformatics promise a fundamental new understanding and order for yeast biology.

“This is all about weaponry,” explains Carroll, noting that Hittinger, in addition to possessing “great benchtop savvy and skill,” has armed himself remarkably well to exploit yeast genetics through the mutually beneficial prisms of molecular biology, evolutionary biology and bioinformatics (which harnesses computers to help make sense of the bumper harvests of data). “He has a determination and resolve to get the answer to any important question— whatever it takes,” says Carroll.

The big questions on the table for Hittinger and others include ferreting out “the genetic factors that drive species diversification and generate biodiversity,” and weaving that granular understanding into the larger fabric of biology. Because the functional qualities of all the various yeast species differ in order for the microbe to thrive in the many different environments it inhabits, the genetic code that underpins their different physiological and metabolic features varies accordingly.

In short, it takes a diversity of talents to inhabit every major terrestrial and aquatic environment the world has to offer. Species that thrive in South American tree galls and species that eke out a living on human skin require different skill sets in order to cope with vastly different environments and utilize different resources. Each of those skills is determined by the organism’s genetic makeup, and as scientists discover and extract the lode of genomic data found in new species discovered in the wild, new and potentially useful genetic information and metabolic qualities will come to light.

These are big, basic biological questions. But their answers promise far more than simply satisfying scientific curiosity. Yeasts are big business. They are medically and industrially important. The secrets they give up will, without a doubt, amplify our ability to produce food, medicine and industrial biochemicals.

To lay the groundwork, Hittinger and an international collaboration of yeast biologists are setting out, with support from the National Science Foundation (NSF), to map the genetic basis of metabolic diversity by sequencing the genomes of the 1,000 or so known species of yeast in the subphylum that includes Saccharomyces. Three hundred times smaller than the human genome, a typical yeast genome consists of 16 linear chromosomes and, roughly, 6,000 genes and 12 million letters of DNA.

“This is the best possible time to be a yeast biologist,” avers Hittinger. “Our collections have been vastly improved, and we can sequence genomes a hundred at a time. The important thing to know is that yeast is not just one organism or one species. There are thousands of yeasts, and they each have their own evolutionary history.”

Acquiring new species from the wild and sequencing their genomes will enable Hittinger and his colleagues to construct an accurate yeast family tree.

“If we don’t understand what’s out there and how they evolved, we’re notgoing to understand how to make use of them,” Hittinger notes. “Now, we can rip ’em open, get a peek at their genomes and see what the differences are and how they’ve changed over time.”

Thus stalking new strains and species of yeast in the wild is an essential part of the program, according to Hittinger, who routinely dispatches students, including undergraduates, to seek out new yeasts in nature. Half of all the known species of yeast have been described scientifically only within the past 15 years, meaning scientists have only a limited understanding of the world’s yeast diversity.

“Until recently, most strain collections have been paltry and biased towards domesticated strains,” says Hittinger. “If we can expand our understanding of the wild relatives, we can use them as an evolutionary model.Yeasts have a much less welldeveloped history in ecology and natural history.”

A recent yeast hunting excursion in Wisconsin by one of Hittinger’s students yielded three strains of the same S. eubayanus lager yeast parent found in tree galls in South America. Discovered near Sheboygan, the yeast has been cultured in Hittinger’s lab and samples have been provided to CALS food science professor James Steele, whose group is setting up a new comprehensive program in fermentation science and, with the help of a gift from Miller-Coors, a new pilot brewery lab in Babcock Hall. (Steele is also looking to support other fermented beverages in Wisconsin—namely, wine and cider— in both production and education. See sidebar on page 20.)

“We grew up a few hundred billion cells, gave them to Jim Steele to brew beer, and we’re eagerly awaiting the results,” says Hittinger, who explains that another focus of his lab is making interspecies hybrids, such as the lager hybrid. “Now that we’ve identified the wild species, we can make crosses in the lab to make hybrids that produce flavors people are interested in.”

In the food science realm, says Steele, yeast research is focused on the functional characteristics—fermentation qualities, sugar utilization, flavors—of a particular strain of yeast. “How does microbial physiology link to flavor in fermented beverages?” he asks.

Saccharomyces strains are the workhorse and best-known yeasts, including many of the most medically and biotechnologically important. With the $2 million award from NSF,
Hittinger and his colleagues will use the genomes to develop a robust taxonomy of important yeasts and look for the genetic footprints that give rise to yeast biodiversity, an  evolutionary history of their metabolic, ecological and pathogenic qualities. Such an understanding will elevate yeast to a new plane as a model and will undoubtedly serve as the basis of valuable new technologies.

Hittinger cautions, however, that sequencing yeast genomes is only a start: “We can very easily read gene sequences, but we don’t yet know how to interpret them fully. We will need to read those bases and make functional predictions” to extend both the knowledge of yeast biology and their potential use in industry.

“But if it weren’t for that natural diversity, we wouldn’t be able to enjoy Belgian beers,” says Hittinger, referencing the gifts conferred by different yeasts and their varied genetic underpinnings, resulting in the different flavors of ales, lagers and Belgians.

One of the central metabolic qualities of the familiar yeasts, of course, is their ability to ferment. Put simply, fermentation is a process by which cells partially oxidize or burn sugar. Among yeasts, the propensity to ferment in the presence of oxygen has evolved only in Saccharomyces species and a few others.

“To make a living using this process, you have to be a glucose hog,” says Hittinger. “But you don’t burn it all the way. You leave some energy on the table. Ethanol burns because it is unoxidized fuel.”

Different kinds of cells can perform fermentation if they become oxygenstarved Human cells, for example, ferment when starved of oxygen, causing painful muscle cramps. Given enough sugar, cancer cells can ferment, and do so to survive in oxygen-poor environments.

Indeed, Hittinger’s research on the cellular resemblance between Saccharomyces yeasts and cancer cells (for which he recently was named a Pew Biomedical Scholar) focuses on identifying which steps in yeast evolution were key to making the transition from respiratory to fermentative metabolic activity, as well as the sequence of those evolutionary events.

“Armed with that information, we should be able to shed some light on how cancer cells make that same transition over an individual’s lifetime,” says Hittinger.

Genes, Hittinger knows, hold the secrets to the functional qualities of yeast. Those microbial secrets, in turn, promise us food, fuel, pharmaceuticals— and, of course, beer. Like bread and wine, the gift of lager is no small thing. Who knows what other gifts, large and small, may lurk in the genes of these microorganisms?


Headed into the wild? If so, you could help Chris Todd Hittinger’s team identify new yeast species and strains. To learn more, visit http://go.wisc.edu/wildyeast

To watch an interview with Chris Todd Hittinger, visit http://go.wisc.edu/hittingerinterview


Brewing Beer-6005

Food science professor James Steele (left) and students are creating a red lager to be brewed by the Wisconsin Brewing Company. Steele and colleagues are launching a fermented foods and beverages program to take research and teaching to the next level.

“Farm to Glass” and More: Fermenting a Growth Industry

We all know Wisconsin as the land of beer and cheese. But in the not too distant future, Wisconsin may also become famous for other fermented products, notably wine and cider, thanks to growing public taste for those products and a blossoming wine- and cider-making culture in the Badger State.

Wisconsin now has about 110 wineries—up from 13 in 2000—and has been adding around a dozen new ones each year in recent years. Many of these operations could use some help, which is on the way in the form of a newly appointed CALS-based outreach specialist whose job is to support the state’s wine and hard apple cider industry.

Leaders of the Wisconsin Grape Growers Association, the Wisconsin Vintners Association and the Wisconsin Winery Association worked with CALS faculty in food science and horticulture to apply for a Specialty Crop Block Grant to support the position through the Wisconsin Department of Agriculture, Trade and Consumer Protection, with the associations providing matching funds. The specialist is scheduled to start working in early 2015.

The position is part of a larger effort to boost fermentation in Wisconsin. CALS food science professor James Steele and his colleagues are laying the groundwork for a comprehensive fermented foods and beverages program through the Department of Food Science—a program that will take to the next level much of the research and teaching the department has been building on for decades.

Already the program is bearing fruit—or, one might more literally say, “bearing beer.” Over the spring 2015 semester, students participating in Steele’s Fermented Foods & Beverages Laboratory will create and develop a new red lager recipe to be brewed by the Wisconsin Brewing Company and sold at the Memorial Union.

A central goal of the program, Steele explains, is to help improve the quality of fermented food and beverage products. As such, the functional roles played by yeast to influence such characteristics as flavor, color and other attributes will be very much in the spotlight.

“Yeast is a key player, beyond the shadow of a doubt,” says Steele. “It is extremely important, but from a food science perspective, it hasn’t gotten a lot of attention.”

With the help of yeast researchers such as Chris Todd Hittinger and his genetics colleague Audrey Gasch, Steele hopes to create an environment where the food science nuances of fermentation are teased out to the benefit of both growers and the producers of fermented foods and beverages.

The basic fermentation characteristics of various yeast strains are of interest, according to Steele: “For example, how does microbial physiology link to flavor in fermented beverages? These collaborations give us opportunities to look for new strains or develop new strains that could allow for the production of beverages with different flavors. And what we learn in one industry, we can apply to another.”

Looking for “Hotspots”

In their quest to make cellulosic biofuel a viable energy option, many researchers are looking to marginal lands—those unsuitable for growing food—as potential real estate for bioenergy crops.

But what do farmers think of that? Brad Barham, a CALS/UW-Extension professor of agricultural and applied economics and a researcher with the Great Lakes Bioenergy Research Center (GLBRC), took the logical next step and asked them.

Fewer than 30 percent were willing to grow nonedible cellulosic biofuel feedstocks—such as perennial grasses and short-rotation trees—on their marginal lands for a range of prices, Barham and his team found after analyzing responses from 300 farmers in southwestern Wisconsin.

“Previous work in the area of marginal lands for bioenergy has been based primarily on the landscape’s suitability, without much research on its economic viability,” says Barham, who sent out the survey in 2011. “What’s in play is how much farmers are willing to change their land-use behavior.”

Barham’s results are a testament to the complex reality of implementing commercial cellulosic biofuel systems. Despite the minority of positive responses, researchers found that there were some clusters—or “hotspots”—of farmers who showed favorable attitudes toward use of marginal land for bioenergy.

These hotspots could be a window of opportunity for bioenergy researchers since they indicate areas where feedstocks could be grown more continuously.

“People envision bioenergy crops being blanketed across the landscape,” says Barham, “but if it’s five percent of the crops being harvested from this farm here, and 10 percent from that farm there, it’s going to be too costly to collect and aggregate the biomass relative to the value of the energy you get from it.

“If we want concentrated bioenergy production, that means looking for hotspots where people have favorable attitudes toward crops that can improve the environmental effects associated with energy decisions,” Barham notes.

CALS agronomy professor Randy Jackson is also interested in the idea of bioenergy hotspots. Jackson, who co-leads the GLBRC’s area of research focusing on sustainability, says that just because lands are too wet, too rocky or too eroded to farm traditionally doesn’t mean they aren’t valuable.

“The first thing we can say about marginal lands is that ‘marginal’ is a relative term,” says Jackson. Such lands have a social as well as a biophysical definition. “This land is where the owners like to hunt, for example.”

The goal of GLBRC researchers like Barham and Jackson is to integrate the environmental impacts of different cropping systems with economic forces and social drivers.

The environmental benefits of cellulosic biofuel feedstocks such as perennial grasses are significant. In addition to providing a versatile starting material for ethanol and other advanced biofuels, grasses do not compete with food crops and require little or no fertilizer or pesticides. Unlike annual crops like corn, which must be replanted each year, perennials can remain in the soil for more than a decade, conferring important ecosystem services like erosion protection and wildlife habitat.

The ecosystem services, bioenergy potential and social values that influence how we utilize and define marginal land make it difficult to predict the outcomes of planting one type of crop versus another. To tackle that problem, Jackson is working with other UW–Madison experts who are developing computer-based simulation tools in projects funded by the GLBRC and a Sun Grant from the U.S. Department of Energy.

Jackson hopes that these modeling tools will help researchers pinpoint where farmer willingness hotspots overlap with regions that could benefit disproportionately from the ecosystem services that perennial bioenergy feedstocks have to offer.

“These models will include data layers for geography, crop yield, land use, carbon sequestration and farmer willingness to participate,” says Jackson. “There could be as many as 40 data layers feeding into these models so that you can see what would happen to each variable if, say, you were to plant the entire landscape with switchgrass.”

Discovery Under Way

What will be the next oil?

That’s a frequent question raised about the future of energy—and not a surprising one considering the dominant role that that single fuel source has played in filling our energy needs.

While we still are searching for the answers to our energy future, one thing seems clear—there probably won’t be one next big thing, one dominant fuel source that will take the place of oil.

Which brings me to the topic of this issue: bioenergy. In 2007 CALS was awarded an initial $125 million from DOE—the largest federal grant ever received by CALS—to come up with new ways of drawing energy from plants. And so we embarked on a scientific endeavor that ranks as one of humankind’s biggest when we consider what we might gain: more ways to free ourselves from dependency on fossil fuels.

The discoveries emerging from these efforts are likely to benefit farmers, businesses and the overall economy in the entire state and region.

While some may have hoped that by this point we’d be tanking up with cellulosic ethanol, anyone familiar with the challenges recognized that after three and a half years, we’d just be warming up.

In fact, we’ve done that and more. As the stories in this issue show—and as an illustration on page 20 offers at a glance—Tim Donohue and his colleagues at the Great Lakes Bioenergy Research Center (GLBRC) have built a research pipeline that already has produced some promising discoveries and is poised to deliver more.

Hundreds of scientists are blazing trails in everything from sustainability—learning how biofuels will affect the environment in the long run—to fundamental research about cell wall growth and interactions with microbes. The GLBRC has strengthened connections with institutions across campus—for example, with the College of Engineering, where researchers are engine-testing biofuels—and across Lake Michigan, working in close cooperation with our partners at Michigan State University. Beyond college campuses, the discoveries emerging from these efforts are likely to benefit farmers, businesses and the overall economy in the entire state and region.

We do not yet know the exact role biofuel will play in the mix of renewable sources that will comprise our energy future. Time and more discovery will tell. We do know that the GLBRC is off to a promising start.

Where Are We Now?

TIM DONOHUE HAS SPENT THE LAST FOUR YEARS BUILDING A PIPELINE—but not the kind that springs to mind when we think of fuel.

The professor of bacteriology heads the CALS-led Great Lakes Bioenergy Research Center (GLBRC), founded with $142 million from the U.S. Department of Energy and a groundbreaking charge—to create the next generation of biofuels by harnessing renewable energy from the nonfood plants that are so plentiful all around us: grasses, trees and crop residues.

“We need to create liquid transportation fuels that are more cost-effective, more sustainable and won’t compromise the Earth or our quality of life,” says Donohue. “We’re in the middle of developing ways to generate these new fuels that are essential for powering our daily lives.”

With Michigan State University (MSU) as UW–Madison’s major partner, Donohue has assembled a team that now includes more than 400 researchers and staff and an additional nine member institutions. The effort spans two countries, 11 states and more than 60 individual lab and field facilities.

That’s a lot of brainpower. But the magnitude of the effort is commensurate with the task at hand, Donohue notes.

“We need to be considering everything from roots in the ground to what’s coming out of the nozzle,” Donohue says. “Without such a holistic approach, we won’t be able to demonstrate that this technology is feasible or see the weak spots where we can make improvements.”

What GLBRC has built is a research pipeline, a process that considers all factors that go into developing and implementing cellulosic biofuels—from creating sustainable agricultural landscapes and building better bioenergy crops to innovations in plant biomass processing and converting plant sugars into fuels.

While the promise of creating sustainable plant-based fuels isn’t new, the level of public investment needed to tackle this challenge has only recently emerged. According to the International Energy Agency, the United States leads world spending on biofuels public research, development and demonstration projects, investing $189 million in 2010 alone.

“By relying on fossils fuels, we’re living on energy that arrived on Earth many millions of years ago,” says Steve Slater, GLBRC’s scientific programs manager. “In order to reach a sustainable energy economy, we need to learn to live on the energy that arrives from the sun today. There’s a lot of that solar energy held within plant biomass, if we can figure out how to sustainably convert it to liquid fuels.”

Four years into its five-year grant, GLBRC has made some significant breakthroughs along the research pipeline. Here are some major points of interest.

First Stop: PLANTS

At agricultural research stations in Wisconsin and Michigan, GLBRC researchers tend to tall stands of such biofuel crops as switchgrass and miscanthus, measuring above-ground traits like crop yield and digging down in the dirt to monitor soil microbes and water movement. Sophisticated instruments measure greenhouse gases such as carbon dioxide and nitrous oxide. Researchers count birds and insects to measure biodiversity and use satellite data to capture a watershed-level view of land use patterns.

It’s a lot of information, but each measurement plays a role in determining how these crop contenders would fare as large-scale bioenergy crops.

The leaves and stalks of these potential bioenergy plants are comprised of large quantities of cellulose, the most abundant organic compound on the planet. Cellulose is a polysaccharide, a long chain of tightly linked sugar subunits that must be broken down into simple sugars before they can be processed into biofuel. That alone is difficult—but to make the process even harder, much of a plant’s cellulose is locked within cell walls that form a tough, protective barrier. Breaking past the walls, using enzymes or chemicals to do so, is one of the biggest challenges in creating economically viable cellulosic biofuels.

Plant cell wall structures have evolved over time to fight off pests and disease. The more scientists understand about how the walls are created, the easier it will be to break them apart. DNA sequencing capacity provided by the Department of Energy (DOE) Joint Genome Institute allows plant breeders access to genetic and genomic data that provide clues about how those cell wall layers are built.

While determining the best genetic traits for bioenergy crops is a long-range goal, GLBRC plant researchers already have made important headway when it comes to tackling lignin, one of the toughest compounds that make up plant cell walls. Researchers hope to take it apart to get at the cellulose locked inside and convert small pieces of lignin into valuable co-products. CALS biochemistry professor John Ralph and his team have identified a gene that would allow easily breakable bonds to be incorporated into plant cell walls. They’re calling this new technology Zip-Lignin™ for its ability to break apart—or unzip—the lignin within. By getting lignin out of the way, biomass processing could be completed at lower temperatures. And lower temperatures mean lower overall costs.

And on another track, GLBRC researchers at MSU have located an enzyme that creates a plant oil with unique biodiesel-like properties. Now they’re working to encourage plants to produce more of that oil, which could be used directly as a “drop-in” or ready-to-use diesel replacement.

Cash Crop Biomass

WISCONSIN FARMERS have been growing biomass for generations, says Kevin Shinners. They just have a different name for it.

“Biomass is really just poor-quality forage,” says the CALS agricultural engineer. “We allow it to get very mature and it’s really high in fiber, so it doesn’t make very good animal feed, but it
makes great biomass.”

And Wisconsin farmers have a leg up in the business of producing biomass, says Shinners, a specialist in forage systems who branched out into bioenergy crops about 10 years ago.

“We have all of the tools to harvest and handle and process it. And an added advantage is that when we take biomass off the field, we have new places to put our dairy manure,” he says. “When you take corn stover off the field, you’re
removing nutrients that you need for next year’s crop. A Wisconsin farmer can apply manure, while an Illinois farmer may have to go out and buy fertilizer.”

Wisconsin also is rich in off-farm resources. The state’s custom harvesters are expert at chopping stalks and grass, and biomass could fit nicely into their schedule. After they finish chopping corn silage in September, crews could move on to corn stover or switchgrass in October and November, spreading fixed costs over more acres and keeping employees working longer.

In fact, under some business models, farmers might job out most of their biomass crop production. If the crop is a perennial, such as switchgrass, the farmer may spend more time in front of the computer and on the phone than out in the field. “Once the crop is established, he’ll manage fertilization and weed control through an agronomic service, cutting and removal through a custom harvester and marketing through a biomass aggregator,” Shinners says.

But even though Wisconsin farmers may be very much at home with the types of crops involved and the mechanics of producing them, they’ll be on less familiar ground when it comes to marketing, Shinners notes.

“If you’re a cash crop farmer, you’re used to marketing your corn and beans through multiple paths, selling some out of the field, storing some, selling futures, to optimize what you earn on an annual basis,” he says. “For biomass, you’ll have to change your mindset.

“If a firm builds a large cellulosic biorefinery here, it will need an absolute dedicated supply,” Shinners says. “If half the people in the area decided not to produce biomass one year, that plant would be a dinosaur.” Meaning that a critical mass of local farmers must be willing to lock into a long-term production contract.

The economics of biomass are driven by the fact that, pound for pound, the stuff isn’t worth as much as other crops. Profit margins may be slim, so farmers will need to produce as efficiently as possible.

That’s where Shinners comes in. His research centers on streamlining the harvest and handling a variety of biomass crops, including such perennials as switchgrass and reed canarygrass, and annuals such as sorghum. But his biggest push has been in corn stover—the stalks and leaves and cobs left when the kernels are removed—simply because there’s so much of it.

“There are some 90 million acres of corn being grown in the United States this year, and with the prices we’re seeing, there’s going to be more and more of it grown. If you’re really interested in biomass, it’s right there at our doorstep,” he reasons.

Since profit-minded crop producers aim to make as few trips across the field as possible, Shinners’ first efforts focused on harvesting both corn grain and corn stover in one pass. Essentially, he grafted a forage harvester to the back of a combine and hitched a wagon behind to catch the chopped stover.

This impressive 50-foot train of machinery worked, he says, but handling two crops at the same time slowed down the grain harvest, putting both yield and quality at risk. “That’s even more of an issue these days, when we have seen corn go over $7 per bushel,” he says. “As corn grain increases in value, everything that slows the combine down has a much greater economic cost.”

Shinners is focusing now on a system in which the combine harvests grain and leaves the stover behind in a long, neat row. “A custom harvester could come in behind and chop these windrows and store them for the farmer.”

Since buyers will need year-round deliveries, storing biomass crop until it’s needed is part of the equation. Shinners thinks the best approach is one that dairy farmers use for forage—seal it from the air in long plastic bags or covered bunkers and let it ferment. “We know this from dairying: You can open up a silo bag from two years ago and it’s still good quality,” he says.

That fermented biomass could be good enough to eat—by livestock, at least—which may offer farmers a way to take advantage of the bioenergy market without having to wait for a biomass refinery to be built nearby.

“If we apply amendments like lime right before we store corn stover, the feed value can increase substantially,” says Shinners. “So instead of waiting for somebody to develop a biorefinery in Wisconsin to convert stover to ethanol, why not divert some of the grain normally used to feed cattle toward ethanol production and use the stover to replace the corn as animal feed?”