Summer 1996
U.S. Department of Energy
Bioenergy Feedstock Development Program at
Oak Ridge National Laboratory
Energy Crops Forum was published periodically by the Bioenergy
Feedstock Development Program, Environmental Sciences Division, Oak Ridge
National Laboratory, managed by UT-Battelle, LLC., for the U.S. Department of
Energy under Contract No. DE-AC05-00OR22725.
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Table of Contents
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The Biofuels Feedstock Development Program (BFDP) is a
mission-oriented program of research and analysis whose goal is to develop
environmentally acceptable crops and cropping systems for producing large
quantities of low-cost, high-quality biomass feedstocks. Major funding comes
from the U.S. Department of Energy's (DOE) Office of Transportation
Technologies with additional funding from the Office of Utility Technologies
and the Office of Industrial Technologies. This issue of Energy Crops Forum
highlights the cooperative nature of several of the research efforts.
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Sheryl Martin and Jerry Tuskan
BFDP
Eastern cottonwood (Populus deltoides) is one of the native poplars of
the Southeast. When properly managed, it outproduces all other southeastern
tree species. These trees perform best on deep, fertile sand-loam to clay- loam
soil with high water-holding capacity. On such sites and under proper
management, cottonwood reaches a height of 30 feet in 5 years, becoming
reproductively mature in 6 to 10 years. Currently, however, we lack well-
tested, broadly adapted clones suitable for short-rotation production.
How do you quickly and efficiently develop superior cottonwood clones that are
resistant to disease and perform well across most of the Southeast? BFDP
decided that the most effective way would be a "virtual crop development
center" that coordinated and united the efforts of experts from institutions
across the region.
Dr. Sam Land, Jr., Mississippi State University (MSU), and his co-investigators,
Dr. Mike Stine, Louisiana State University and Dr. Donald Rockwood, University
of Florida, have created just such an initiative for BFDP. Their "virtual crop
development center" includes a host of cooperators across 13 southeastern
states—Boise Cascade Corporation (Louisiana), Champion International Company
(Florida), Northwest Florida Water Management District, Crown Vantage
(Mississippi), International Paper Company (Georgia), Neal Land and Timber
Company (Florida), Packaging Corporation of America (Tennessee), Tennessee
Division of Forestry, Union Camp Corporation (Georgia), and Westvaco (Kentucky
and South Carolina). Their goal is to develop eastern cottonwood (Populus
deltoides) clones suitable for short-rotation woody crops in the
Southeast.
How do you coordinate the efforts of this many people? "Frantically," says Land.
The cooperators locate natural stands, obtain flowers and seeds, and provide
and maintain test sites. Sites for collecting flowers and seeds stands have
already been located throughout the Southeast, and new test sites are being
prepared in west Tennessee, Alabama, northwest Florida, and South Carolina.
Cottonwood flowers are pollinated and the seed is dispersed in a short time
period, typically within 3 or 4 weeks. Seed must be collected prior to seed
shedding so that its origin is known. Spring is also the only time to collect
limbs containing flowers and to bring these limbs back to the laboratory to
make controlled crosses.
When a cooperator reports trees are in flower or that seeds are mature, everyone
must be ready to move. Land contacts his assistants, who are often far away
from MSU. He puts those assistants in touch with the cooperating institution's
field people to coordinate the collection of new material. The field workers
also collect leaf tissue for biotechnology studies at Louisiana State
University.
The work does not end there. Summers are spent maintaining a breeding orchard.
In the winters, cuttings of the selected genotypes from the nursery are
prepared. Throughout the year, everyone is on the lookout for new cottonwood
stands.
Not just any cottonwood will do. Selections are based on relative growth,
apparent disease resistance, and wood specific gravity. The goal is to collect
plant material from a total of 24 areas, four areas in each of six subregions
across the Southeast. The areas coverdifferent river systems and different
parts of river systems and include river bottom lands and upland stream sites.
After selection and collection are completed, the researchers cross- pollinate
selected plants to develop new clones that are genetically suitable for
short-rotation woody crops in the Southeast.
Mississippi State University's Delta Research and Extension Center is the hub of
the cottonwood development effort. This facility was previously used in the
1960s and 1970 by the U.S. Forest Service for cottonwood selection and
improvement. The University of Florida's contribution includes the
establishment and maintenance of a test site and nursery near Tallahassee,
Florida. The industrial cooperators are providing additional test sites.
Louisiana State University will use the leaves collected during seed sampling
to construct gene maps consisting of DNA markers. Relating these markers to
traits of commercial interest, such as growth rate or disease resistance, will
permit early identification of desired traits in offspring, eliminating a long
wait for plants to exhibit superior performance.
Through modern technology and old-fashioned cooperation, the process of tree
improvement, one of the oldest procedures in forestry, is being done faster and
more efficiently by Land and his collaborators across the Southeast in BFDP's
new "virtual crop development center.
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Toby Bradshaw
University of Washington
As intensive culture of Populus species and hybrids becomes increasingly
important as a source of fiber and fuel, breeding strategies must be developed
and implemented to measure, capture, recombine, and fully exploit the abundant
genetic variation still found in natural populations of poplars and
cottonwoods. Adaptation of poplars to the specialized needs of industry is
still in its earliest stages and must take advantage of the best genetic
methods available if rapid, sustained genetic improvement is to be made.
Since 1989, molecular genetic investigations into the basis of Populus stem
growth, form, stress tolerance, and disease resistance have pinpointed regions
of the Populus genome that have significant effects on these traits. In
collaboration with tree physiologists, anatomists, biochemists, and
pathologists, we are at the threshold of a new era in poplar breeding in which
these few key regions of the genome may be manipulated efficiently and then
recombined into novel hybrids with the best characteristics of several Populus
species.
The time is right to capitalize on the insights into poplar breeding that
molecular biology offers. Significant gains can then be expected in both the
near and longer term. For "molecular breeding" to flourish, however, critical
research needs must be addressed. Informative genetic material must be
identified, collected, bred, and tested over a wide range of actual and
potential growing areas. Although previous clonal testing and breeding have
offered useful guidelines, new research will expand the scale of breeding and
testing while increasing their pace and precision through the use of molecular
tools.
Data gathering, storage, retrieval, and analysis must be coordinated to ensure
rapid dissemination of useful information. Stability and continuity of funding
are required to permit the development of highly informative multigeneration
pedigrees within a large mating design, to explore alternative breeding
strategies, and to retain key personnel. At the same time, immediate and
continued production of clones with potential commercial use is needed.
Each of these research needs will require extensive collaborative efforts on the
part of industry, government, and academic institutions. In order to coordinate
research efforts among these organizations, the Poplar Molecular Genetics
Cooperative (PMGC) was formed in March, 1995. PMGC's goals are to increase
understanding of the molecular genetic mechanisms causing variation in
productivity and quality traits in hybrid poplar and to use research results to
accelerate progress in poplar breeding.
The key to sustained genetic improvement in Populus is a detailed
understanding of the genetic architecture of important phenotypic traits (e.g.,
stem volume, wood quality, disease resistance). Such an understanding will
accelerate progress in breeding by providing improved methods to identify
superior clones for immediate commercial use and parents for the next
generation of hybrids. Crucial questions to be addressed include:
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Which regions of the Populus
genome contribute to heterosis (hybrid vigor) in interspecific hybrids?
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Which regions of the genome are responsible for variation in clonal performance
and breeding value?
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Is clonal performance a good predictor of breeding value?
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What useful traits do each of several Populus
species bring to their hybrid offspring?
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How can clones be genetically tailored to match specific sites and
silvicultural regimes, as well as manufacturing processes and products?
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Which breeding and selection system(s) will maximize genetic gain in the near
and long terms?
These questions have no simple or obvious answers, but with molecular tools they
are now approachable in a reasonable time frame.
In the first year of PMGC's existence, breeding methods suitable for handling
large numbers of crosses were developed and implemented. Especially significant
were techniques that enabled researchers to isolate female branches from
unwanted pollen and to rescue immature embryos by in vitro culture. More
than 1800 new hybrid seedlings were planted at the Washington State University
Farm 5 in Puyallup, Washington. The industry demand for new genetic material
was very strong; more than 10,000 cuttings were requested from the first-year
PMGC seedlings for immediate placement in company-sponsored clonal performance
trials.
PMGC members support the cooperative with annual dues of $12,500 and with
in-kind contributions of land, equipment, personnel, and expertise. PMGC is
also working with nonmembers who provide additional breeding materials and
technical advice and support. Thus, monies from DOE and BFDP are leveraged
significantly by industry and academic institutions around the world. The
current PMGC membership includes Alberta-Pacific Forest Industries; Boise
Cascade Corporation; Georgia-Pacific West, Inc.; Inland Empire Paper Company;
James River Corporation; MacMillan Bloedel Timberlands, Inc.; Nippon Paper
Industries Company, Ltd.; Potlatch Corporation; Scott Paper, Ltd.; Union Camp
Corporation; Weyerhaeuser Company; British Columbia Ministry of Forests
Research Branch; Washington State University; the University of Washington;
and, of course, DOE and BFDP.
For additional information, contact Toby Bradshaw at the Center for Urban
Horticulture, Box 354115, University of Washington, Seattle, WA 98195-4115,
(206) 616-1796 (voice), (206) 616-1826 (FAX), or toby@u.washington.edu (email).
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Lee Gunter, Jerry Tuskan, and Stan Wullschleger
BFDP
Crop breeders have long sought to identify genetically controlled traits that
are associated with superior plant growth and dependable productivity and that
can be used as indicators in the selection of higher yielding cultivars. Unlike
many forage and cereal crops, which have been in domestication for centuries,
relatively little is known about the genetics of switchgrass, BFDP's model
herbaceous energy crop. Such a deficiency makes development of superior
cultivars difficult. Across its wide native geographic range, switchgrass has
evolved into upland and lowland ecotypes (locally adapted populations) that
vary in morphology and physiology. The considerable variation in switchgrass
will provide plant breeders with the means to increase biomass yields.
BFDP hopes to simplify the assessment of switchgrass germplasm by using a
technique that reveals variation in randomly distributed genomic DNA segments.
It is now possible for geneticists and breeders to routinely extract DNA from
small amounts of leaf tissue and make many copies using the polymerase chain
reaction (PCR), a process that mimics the enzymatic amplification of DNA within
a single cell. These amplified segments can serve as markers for attributes
such as disease resistance or drought tolerance that may not be apparent at the
seedling stage. In such cases, the marker is assumed to be genetically linked
to the trait. Often, however, these PCR-based markers (known as random
amplified polymorphic DNA or RAPD markers) act as simple genetic "fingerprints"
and are useful tools to enhance our ability to perceive differences between
very similar individuals, cultivars, varieties, or breeding lines.
One of the conditions that makes successful switchgrass breeding difficult is
variation in chromosome numbers, because reliable inheritance of
genetically-controlled traits is based on stable chromosome arrangements across
generations. Switchgrass ecotypes are known to have multiple copies of a basic
chromosome number: typically 4X in lowlands and 6X to 8X in uplands, according
to observations made by Charles Taliaferro and Andy Hopkins at Oklahoma State
University1.
Our first objective was to search for markers that could be used to
differentiate switchgrass cultivars on the basis of chromosome number or
ecotype. Using RAPD primers in the PCR amplification process, we detected a
single marker in plants with higher chromosome numbers2. The marker
is present in 8X cultivars (e.g., 'Shelter' and 'Trailblazer') but is absent
from 4X cultivars (e.g., 'Alamo' and 'Kanlow'). This finding is in accord with
the work of University of Nebraska agronomist Ken Vogel, who developed markers
that distinguish 4X and 8X cultivars based on chloroplast DNA restriction
fragment length polymorphisms (RFLPs3). We anticipate that these research
results will enable breeders to make a quick assessment of native germ plasm
collections for potential genetic compatibility without resorting to
labor-intensive chromosome counts.
Because chromosome number is not an absolute predictor of ecotype in
switchgrass, we are assembling a set of diagnostic markers for estimating
relationships between new germ plasm and released cultivars by comparing
standard morphological descriptors to molecular markers. While both diagnostic
markers and morphological descriptors are useful, preliminary evidence suggests
that molecular markers, which are more abundant, are better predictors when
cultivars are morphologically similar. The ability to make such distinctions
will facilitate the organization of switchgrass germ plasm breeding
populations.
The use of molecular markers in switchgrass research seems promising. We hope
the application of these methods and others will encourage collaboration among
researchers to increase our understanding of switchgrass biology, thus
overcoming obstacles in the development of superior cultivars.
References
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A.A. Hopkins and C.M. Taliaferro. Chromosome number and nuclear DNA content of
several switchgrass populations. Crop Science
(in press).
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L.E. Gunter, G.A. Tuskan, and S.D. Wullschleger. 1996. Diversity among
populations of switchgrass based on RAPD markers. Crop Science
36:1017–1022.
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S.J. Hultquist, K.P. Vogel, D.J. Lee, K. Arumuganathan, and S. Kaeppler. 1996.
Chloroplast DNA and nuclear DNA content variations among cultivars of
switchgrass, Panicum virgatum L. Crop Science 36:1049–1052.
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Anderson, I.C., D.R. Buxton, and J.A. Hallam. 1996. Selection of Herbaceous
Energy Crops for the Western Corn Belt. Final Report Part I: Agronomic Aspects.
ORNL/Sub/88-SC264/P1. Oak Ridge National Laboratory, Oak Ridge, Tennessee.
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Anderson, I.C., D.R. Buxton, and J.A. Hallam. 1996. Selection of Herbaceous
Energy Crops for the Western Corn Belt. Final Report Part II: Economic
Potentials. ORNL/Sub/88-SC264/P2. Oak Ridge National Laboratory, Oak Ridge,
Tennessee.
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Bongarten, B.C., and S.A. Merkle. 1996. Optimizing Energy Yields in Black
Locust Through Genetic Selection: Final Report September 30, 1994.
ORNL/Sub/86-95907/4. Oak Ridge National Laboratory, Oak Ridge, Tennessee.
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Dickmann, D.I., K.S. Pregitzer, and P.V. Nguyen. 1996. Net Assimilation and
Photosynthate Allocation of Populus
Clones Grown under Short-Rotation Intensive Culture: Physiological and Genetic
Responses Regulating Yield. ORNL/Sub/86-95903/4. Oak Ridge National Laboratory,
Oak Ridge, Tennessee.
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Meyer, D.W., W.E. Norby, D.O. Erickson, and R.G. Johnson. 1996. Evaluation of
Herbaceous Biomass Crops in the Northern Great Plains: Final Report August 31,
1994. ORNL/Sub/88-SB844/2. Oak Ridge National Laboratory, Oak Ridge, Tennessee.
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Stettler, R.F., T.M. Hinckley, H.D. Bradshaw, Jr., and P.E. Heilman. 1996.
Genetic Improvement and Evaluation of Black Cottonwood for Short-Rotation
Biomass Production: Final Report 1987 to 1992. ORNL/Sub/83-43382/8. Oak Ridge
National Laboratory, Oak Ridge, Tennessee.
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