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Short Rotation Woody Crops Research |
Objective: Establish experimental plantings of 39 willow and poplar clones to determine clonal and environmental effects on growth, understand biomass partitioning and nutrient use efficiency, and understand pest utilization of clones. Maintain and expand cutting orchards of genetically improved willow clones in preparation for commercial-scale trials.
Approach/Background: The State University of New York College of Environmental Science and Forestry (SUNY ESF) is expanding its willow genetic improvement program to meet future needs for genetically improved willow clones. Basic physiological information on clones currently being propagated and tested on a large scale, and new clones being propagated in preparation for testing, is necessary to assist in clonal selection and the overall genetic improvement effort. Large genotype-by- environment interaction is a problem facing willow breeders, but the physiological basis of these interactions are poorly understood.
Cutting orchards were established at Syracuse (19 willow clones, 1 hybrid poplar clone, 60 cuttings per clone), and Tully, NY (16 willow clones, 60-200 cuttings per clone) during 1993 with funding from USDOE-ORNL. A willow cutting orchard was established during 1995 in cooperation with the New York State Energy Research and Development Authority and the New York State Department of Environmental Conservation (NYDEC) at the NYDEC Saratoga Tree Nursery, Saratoga, NY, and expanded during both 1996 and 1997 with support from the USDA Cooperative States Research and Extension Service. The University of Toronto (UT) established a cutting orchard at Orono, Ontario during 1992 with 18 willow clones and 600-1200 cuttings per clone.
Status/Accomplishments: A trial was hand-planted at Tully, NY containing 32 willow and 7 hybrid poplar clones at 0.6 by 0.9 m spacing during May, 1997. Site preparation for this trial was completed during fall 1996, consisting of contact herbicide application, plowing, disking, and capping with oxyfluorfen preemergent herbicide (Goal 1.6e) at the 2.24 kg ha-1 rate. The experimental design was a randomized complete block with four replications. Experimental plots contained 36 trees. A sister study was planted during June 1997 in Rhinelander, WI in cooperation with the USDA Forest Service. In the Tully trial, time of budbreak, and survival were measured during spring, 1997. Weekly measurements of stem diameter, tree height, and water stress, and biweekly measurements of numbers of leaves, leaf area, and pests were completed during summer 1997. Meteorological data were collected continuously since the trees were planted.
Approximately 300,000 cuttings were produced by the Saratoga tree nursery during winter 1996-1997, and conservatively, this number could have been as high as 400,000 but we did not have a need for the cuttings during 1997. UT and SUNY ESF produced approximately 200,000 and 150,000 cuttings, respectively, for planting in spring 1997, and UT could have produced far more. The willow cutting orchard established during 1993 at Tully, NY was irrigated and fertilized during the 1997 growing season. A new cutting orchard containing 95 new willow clones collected from wild stands (50-100 cuttings per clone) was planted during 1997 at Tully. The Saratoga cutting orchard was expanded by approximately 18,000 cuttings during spring 1997. The future of the Orono, Ontario cutting orchard is uncertain because the Ontario Ministry of Natural Resources decommissioned the Orono Nursery. Currently the cutting orchard is intact but not being irrigated.
Objectives: The long-term goal of this research is to support the development of biomass production systems for energy conversion by providing a more fundamental understanding of drought tolerance mechanisms that can be exploited to achieve high productivity on potentially stressful sites. The primary objectives of the research are to determine 1) the extent of drought tolerance of poplar in currently available germplasm, 2) QTL for drought tolerance, analyzed by using structured multiple-generation pedigrees, and 3) the extent that drought tolerance can be increased through selective breeding.
Approach/Background: A recently-completed Cooperative Research and Development Agreement (CRADA) between Oak Ridge National Lab (ORNL) and Boise Cascade Corp. (BCC) provided a validation of the role that low osmotic potential and osmotic adjustment have in determining drought tolerance of poplar clones in the field. Follow-on collaborative research in the Pacific Northwest between ORNL, BCC, and University of Washington (UW) is focussing on the advanced-generation poplar pedigrees and clones from diverse crosses generated by UW and established and managed by BCC at Ice Harbor Fiber Farms, near Pasco, WA. Pure clones and hybrids of P. trichocarpa (T), P. deltoides (D), and P. nigra (N) are being studied. The research is determining osmotic potential at full turgor (Ypo) and identifying the most promising of the drought tolerant, high productivity F1 clones for selective breeding to enhance tolerance. Simultaneous analysis of clones from a structured, outbred pedigree for Ypo and solutes that consititute Ypo, will permit marker identification.
Status/Accomplishments: Over 90 pure and hybrid poplar clones were screened for variation in Ypo. Plants were in their first or second growing season, at BCC.'s Ice Harbor Fibre Farm, near Pasco, Wa. Of trees in their first growing season, the pure Populus deltoides clones averaged -1.70 MPa, which was more than 0.20 MPa lower than the mean for P. maximowiczii (-1.48 MPa) and P. trichocarpa (-1.46 MPa) clones. Of the trees in their second growing season, the TxD hybrids typically had the lowest Ypo (-1.67 MPa; 1/3 of the clones <-1.75 MPa), followed by P. trichocarpa (-1.52 MPa), TDxD (-1.50 MPa), and TxN (-1.47 MPa) hybrids. Although pure P. deltoides clones have the lowest Ypo observed (-1.92 MPa), backcrossing D with TD, overall, does not lower Ypo of the resulting progeny below that observed for P. trichocarpa.
Summary Date: October 1997
Objectives: Use molecular markers and genome maps to pinpoint quantitative trait loci (QTLs) controlling tree growth, wood properties, stress tolerance, and disease resistance to increase understanding of the molecular genetic mechanisms that cause variation in productivity and quality traits in hybrid poplar. Use research results to accelerate progress in poplar breeding.
Approach/Background: The key to sustained genetic improvement in Populus is a detailed understanding of the genetic architecture of important phenotypic traits such as stem volume, wood quality, and 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.
Genetic maps composed of molecular markers provide a powerful tool for studying the inheritance of biomass yield and quality traits. Previous results from QTL mapping in three-generation hybrid poplar pedigrees indicate that there are chromosomal regions with large effects on growth, form, stress tolerance, and disease resistance. These results are being confirmed and extended by creating new QTL mapping pedigrees of very large family size, developing 'universal' genetic maps for these families, and testing a wider variety of traits, including the chemical and physical components of the woody biomass.
The PMGC breeding program is designed to produce genetically-informative pedigrees, but also is likely to generate commercially-valuable hybrid poplar clones for industry in the Pacific Northwest, upper Midwest, and Southeast.
Status/Accomplishments: More than 200 full-sib hybrid poplar families and 15,000 seedlings have been produced for molecular genetic analysis, field testing, and potential commercial deployment. More than 10,000 cuttings were supplied to industrial partners this year alone.
Individual F1 (P. trichocarpa x P. deltoides; TxD) and backcross (TDxD) hybrid families with as many as 2000 offspring will allow very fine scale genetic mapping of sub-chromosomal regions conferring beneficial traits, such as resistance to disease. Fine scale genetic mapping will be followed by physical mapping and cloning of disease resistance genes. The first two targets of this fine structure mapping are resistance genes for Melampsora leaf rust and Septoria stem canker, the two most economically- important disease of poplars in North America.
Development of microsatellite markers for construction of a 'universal' poplar genetic map has begun. Markers with the CTT trinucleotide repeat have proven to be highly polymorphic and useful across a wide range of Populus species and hybrids.
PMGC membership has grown to include 12 forest products companies, the B.C. Ministry of Forests, and two universities, in addition to the DoE BFDP. The DoE BFDP contribution is leveraged 4:1 by the other PMGC members/partners. PMGC support of 'startup' projects has also led to the acquisition of an additional $187,000 in grants from the USDA NRI Competitive Grants Program, the US Forest Service, and the Consortium for Plant Biotechnology Research.
Summary Date: September 1997
Objective: Develop improved genetic varieties (clones) of eastern cottonwood (Populus deltoides Bartr.) and its hybrids for use in energy and fiber crops throughout the southeastern United States. Research will identify appropriate sources (provenances) and clones for different parts of the region, investigate methods for stimulating early flowering, and determine if Quantitative Trait Loci (QTL's) can be identified for yield traits.
Approach/Background: Task 1--Assemble plant material. The southeastern region of the U.S. (NC- OK southward to FL-TX) will be subdivided into six subregions. Open-pollinated seeds from natural stands in 3 subregions east of the Mississippi River and cuttings from tested clones in the other 3 subregions (southern Mississippi River and west of River) will be collected during the first 3 years (1996-98).
Task 2--Develop multi-generation pedigreed plant material. Crosses among the tested clones will occur during the first 3 years (1996-98) to produce pedigreed material (control-pollinated seeds) for second-generation genetic improvement and for QTL analyses.
Task 3--Test the assembled and developed plant material. Field trials with cuttings from the o.p. and c.p. seedlings will be established on both river-bottom and upland sites in all 6 subregions during the period 1999-2003. Selection of clones in these tests will be based on rapid juvenile growth rate, Melampsora leaf rust resistance, and high wood specific gravity.
Status/Accomplishments: Task 1-- Sixty-seven of 72 desired natural stands have been located, and leaves and open-pollinated seeds were collected from 1-5 mother trees in each of 54 of these stands during April-June 1997. Viability of many of the seed lots was poor, because of immature seed collection or incorrect storage (literature was lacking or incorrect). DNA has been prepared at LSU from the leaf collections to test markers for QTL analyses. Cuttings from 220 tested clones with origins in the 3 western subregions have been planted in nurseries and breeding orchards at Stoneville, MS, Baton Rouge, LA, and Quincy, FL. Of these, 214 clones have at least one surviving ramet at Stoneville.
Task 2-- During February 1997, limbs with reproductive buds were collected from 36 female and 35 male tested clones in the 3 western subregions. Eighty-one of 140 planned crosses were controlled pollinated during March 1997, and seed from 7 of these crosses were collected (without embryo rescue) in June.
Task 3-- Seeds of approximately 40 open-pollinated mother-tree families have been germinated by LSU and seedlings from 14 families are being vegetatively propagated by University of Florida. Six forest industries have agreed to provide test sites.
Summary Date: September 1997
Objective: Our objective is to expand and accelerate poplar breeding and selection in the North Central United States by implementing a regional testing program for Minnesota, Iowa, Wisconsin and Michigan. Tests are established every two years and test clones are evaluated through at least 1/2 rotation age (5 to 6 out of 10 to 12 years). Multiple traits are evaluated simultaneously, including tree height, stem caliper, and resistance to biotic and abiotic stresses. Discriminant function selection indices are developed to increase the probability that crop characteristics are commercially suitable in the aggregate. Collaborators include experts in pathology, entomology, physiology and genetics to provide a multidisciplinary approach to breeding and selection.
Approach/Background: This project provides estimates of genetic and environmental effects on the growth of new hybrid Populus clones. First, we are interested in the magnitude of clonal variation at each location individually, and especially in how new clones perform relative to current commercial standards. Second, we are interested in the stability of clonal performance across locations. Third, we are interested in determining how environmental conditions and the distribution of diseases affects stability. Overall, results of the project should include new clonal selections that equal or exceed the performance of existing standards and also new information on clonal stability that will: 1) allow the delineation of breeding zones in our region and 2) guide funding decisions so that research investment matches crop development needs. We are conducting this research because there have been no regional cooperative tests of newly developed Populus clones in our region in the last 30 years.
Our approach to the research is to conduct common garden experiments where all clones to be tested are planted at each of four locations in Minnesota, Iowa, Wisconsin, and Michigan. We measure tree height, tree diameter, and the incidence of diseases at each location. Data are analyzed using Analyses of Variance and Covariance. We then estimate variances attributable to locations, blocks within locations, clones, the clone x location interaction, and within plot error. Variance and covariance components are used to estimate heritabilities on both an individual location basis and over all locations, clone x location interactions (an indication of stability) and genetic correlations among various plant traits. Variance and covariance components are also used to predict the outcome of various hypothetical multiple trait index selection strategies.
Testing focuses on Eastern cottonwood (Populus deltoides) which has potential as a biofuels crop per se and also has potential value as a parent in interspecific hybridizations. Eastern cottonwood has been selected as the primary species for woody biofuels development in the North Central United States because of its relative resistance to Septoria canker, which limits deployment of many poplar hybrids in the region. Eastern cottonwood is also a native species in the North Central region which may be an important consideration for deployment in disturbed riparian ecosystems where social and environmental concerns weigh heavily. We also test promising inter-specific cottonwood hybrids and some hybrid aspen.
This project operates on a cooperative basis, accomplishing objectives through several organizations including Pope County (Minnesota) Soil and Water Conservation District, the University of Minnesota, Iowa State University, the University of Wisconsin, and Michigan State University. We continue to support our cooperators in the Regional Testing of Populus Clones through Federal Assistance Agreements with North Central Forest Experiment Station (NCFES). Funds received through these Agreements are used to complete site preparation, planting, plantation maintenance and measurements.
Status/Accomplishments: We have established two sets of tests thus far, one in 1995 and one in 1997. Clones were selected from nurseries at Iowa State University and the University of Minnesota (Grand Rapids, Minnesota) for inclusion in the 1995 planting based on age two year growth and resistance to leaf diseases and also on availability of ramets in sufficient numbers to support the planned replication. Overall, we selected 43 clones of P. deltoides, 10 clones of P. deltoides x P. maximowiczii F1 hybrids, two clones of P. deltoides x P. nigra F1 hybrids (including the DN-34 [a.k.a. NC-5326, cv. Eugenii] control), one clone of P. nigra x P. maximowiczii (NM-6 control), and four clones of aspen hybrids. The commercial control clones in the experiment, NM-6 and DN-34, are, respectively, one of the most promising new commercial clones and, currently, the most widely planted commercial clone in the Northeastern US. Tests were established at Westport, Minnesota; Ames, Iowa; Madison, Wisconsin; and East Lansing, Michigan (Figure 1) using 1-0 rooted plants that had been top and root pruned. In each case, the planting design was 10 randomized incomplete blocks with two tree plots.
We reared one-year-old rooted plants for the 1997 planting in the nursery at Grand Rapids, Minnesota. The experimental design in 1997 was changed because evaluation of early data from the 1995 plantings suggested we could obtain 90% of the precision associated with 10 replications by planting only 5 replications. This allowed us to increase the number of clone entries to nearly 90 while holding the overall experimental size unchanged. The 1997 plantings, established adjacent to the 1995 plantings at all locations, contain 70 clones of P. deltoides, 17 clones of P. deltoides x P. maximowiczii F1 hybrids, and the two commercial standards, DN-34 and NM-6.
We have completed analyses of height and stem caliper growth at age 1 year in the 1995 test at all four locations, and of height and stem caliper at age 2 years at the Westport and Ames locations. About 30 experimental clones exceeded the commercial standards in height and caliper at the end of the first year. Clone x location interactions were small compared to clonal effects, indicating that clone performance was stable across the region. However, at the end of the second year we measured substantial dieback among clones of southern Iowa and Illinois origin in the most northerly tests. Clone x environment interactions began to dominate the results suggesting that adaptation to cold winters may severely limit the transfer of select clones, even between breeding programs as geographically proximate as Iowa and Minnesota. We are propagating 14 clones from the 1995 test to support large plot testing in Minnesota based on height, stem caliper and resistance to Septoria canker at the Westport site.
This Project collaborates with the Minnesota Hybrid Poplar Research Cooperative to extend the Regional Field Test project into northern Minnesota. Collaborating institutions include the Natural Resources Research Institute, Duluth, The University of Minnesota - Crookston, and the Agricultural Utilization Research Institute, Crookston. We established additional plantings of the 1995 Regional Test clones at several sites in Minnesota in 1997. In addition, The Minnesota Cooperative supports an extensive breeding program that will contribute new clones to the Regional Field Test Program as early as 1999.
Summary Date: July, 1997
Objective: Select and breed Populus deltoides germplasm with the best potential for biomass production in the region: screen this germplasm for pest resistance, rootability, and initial growth; produce clonal stock of the best selections to support field testing of adaptability and productivity; establish correlations between laboratory, nursery, and field traits; quantify pest impacts; and evaluate novel types of pest resistance.
Approach/Background: In July 1988, this project was started. Emphasis is being placed on selection and breeding for pest resistance, dry weight yield potential, and ease of propagation. Selected parents are bred and natural populations are sampled to produce initial progeny and clonal tests. Promising individuals are placed into one-year, replicated clonal trials in the nursery. Clones that have resistance or good tolerance to Melampsora, Marssonina, and Septoria leaf diseases; resistance to Septoria stem canker; acceptable levels of cottonwood leaf beetle and stem insect damage and above average growth are scaled-up and supplied to the U.S. Forest Service/DOE project on regional testing of new clones.
Basic research is conducted on correlations between nursery and field performance, mechanisms of resistance to Septoria canker (Septoria musiva) and cottonwood leaf beetle (Chrysomela scripta), and the impact of the cottonwood leaf beetle on biomass production.
A number of cooperators provide pollen and flowering branch material, test sites, and collaboration on research questions. During the past two years, we have had significant cooperation from the Iowa DNR Nursery, Boise Cascade, Westvaco, numerous scientists at Iowa State University, colleagues at the universities of Illinois, Minnesota, Mississippi State, Northern Arizona, SUNY, Syracuse, Toronto, Washington, Washington State, Wisconsin, and U.S. Forest Service researchers. Substantial assistance has also come from overseas from Long Ashton Research Station, University of Bristol, UK and INRA, Orleans, Fr.
Status/Accomplishments: In 1996, we provided cuttings for 86 clones to be entered in the second round of regional trials in Iowa, Michigan, Minnesota, and Wisconsin. In other tree improvement work, we continued development of a new breeding techniques for forcing pollen and holding female flowers through to seed maturation, began producing P. deltoides X P. maximowiczii hybrids to study Septoria canker resistance and produce improved clones, and began making P. deltoides X P. nigra and P. maximowiczii X P. nigra crosses to evaluate hybrid vigor using selected parents. From one P. deltoides family, we selected and are propagating two sets of 16 clones each that are segregating for a single gene trait conferring Melampsora rust resistance. We have propagated over 6200 open-pollinated and control- pollinated seedlings to establish new progeny tests in 1997-8.
Our pathology studies clearly indicate that the population of Melampsora medusae has changed in Iowa since 1992. In addition, further evidence supports the hypothesis that a new pathotype of Marssonina brunnea has become established in Iowa. Continued studies on the anatomy of bark tissue in young stems of greenhouse-grown trees indicate that propensity for meristematic activity in the phellogen and parenchyma tissues is a characteristic of Septoria canker resistance. The use of this characteristic, coded with other anatomical and physical features of this young tissue, may lead to early selection criteria for canker resistance.
In our entomology studies, two chemicals (C22-30 alcohols and a-tocopherylquinone) that together induce adult cottonwood leaf beetle feeding were identified from the leaf surfaces of Populus clones. Adult feeding preferences and leaf-surface chemicals were quantified among University of Washington parent, F1, and F2 pedigree material. We found that between 0.2 and 1.1 egg masses per actively growing terminal are sufficient to cause economic injury. Life table studies and surveys of leaf beetle predator activity suggest that first and third generations are more suppressed by natural agents than is the second. The phenology of the cottonwood twig borer (Gypsonoma haimbachiana) in central Iowa was verified; two generations per year occur in Iowa.
Summary Date: September, 1997
Objectives: The objective of this research is to minimize the damage to hybrid poplar plantings caused by Septoria spp. using genetic resistance and somaclonal selection techniques to develop reliable clones. Screening clones for resistance to the canker and leaf diseases caused by Septoria spp. requires a thorough understanding of the population structure, including the pathogenic variation and distribution of the fungi causing these diseases. These are the major topics of this investigation.
Approach/Background: We have used cell and tissue culture techniques to generate a population of somaclonal variants from two hybrid poplar clones that have rapid early growth but are highly susceptible to Septoria canker and leaf spot diseases. These regenerated plants have been selected using a laboratory bioassay to identify individuals with increased disease resistance. Selected resistant somaclones have been clonally propagated and established in replicated field tests to validate the increased resistance expressed by the somaclones under field plantation conditions.
Septoria musiva has severely damaged many plantings of susceptible hybrid poplars in the north central and northeastern United States but has not been found to occur in the Pacific Northwest. S. populicola is common on leaves but does not cause stem cankers on black cottonwoods in the Pacific Northwest and on native balsam poplars elsewhere in North America. The distribution of these fungi and their pathogenic variation is not well understood, which complicates screening poplars for resistance. Over 1000 isolates of Septoria spp. have been collected from various native and hybrid poplars over a wide geographic range in order to examine the genetic variability within populations of this fungus using RAPD markers. Variation in pathogenicity is also being assessed by inoculating a set of poplar host differentials.
Status/Accomplishments: We have under field test several somaclones regenerated from tissue cultures of 2 hybrid poplar clones. These somaclones have previously expressed increased resistance to Septoria leaf spot in laboratory bioassays compared to the donor clone controls. The first field planting was established near Rhinelander, WI in 1986 and consisted of the original plants recovered from tissue culture and non-cultured donor clone controls. Hardwood cuttings taken from trees expressing field resistance were used to establish replicated field trials near Rosemount, MN in 1991 and 1996 to determine the stability of the resistance after conventional clonal propagation. Many of these trees have retained their resistance in these plantings as well. In 1995 a planting was established near Rosemount using somaclones recovered from a different hybrid poplar clone. After 3 years, most of the somaclones selected from the laboratory screening technique are superior to the controls in growth and disease resistance. The mechanisms that are responsible for the somatic variation we have observed are not fully known, however, it is possible that some poplar clones are more unstable than others when passed through a tissue culture cycle and useful traits such as disease resistance may be uncovered using these techniques.
We are continuing our study of the genetic and pathogenic variability of S. musiva and S. populicola isolates that have been collected from over 50 different hybrid and native poplar clones in 8 states and Canada. The fungi vary widely using various morphometric measurements and RAPD markers. Conidial morphology among the species is highly variable and often overlaps between them. It has been possible to partially differentiate among the species using molecular markers. Although S. populicola has not been known to cause cankers in the field, cankers do develop on stems of inoculated plants in the greenhouse. The pathogenic variability and possible host specificity among selected isolates are now being examined using laboratory and greenhouse tests of excised leaves and whole plants.
Various field trials of hybrid poplars in the north central region are continuing to be monitored in order to identify those clones that are best adapted to local growing conditions and that are resistant to the major diseases in the region. In addition, plantings of P. trichocarpa pedigree families are being evaluated for their resistance to local leaf rust and Septoria canker to determine genetic control of resistance to these important pathogens and to learn if clones selected in one region can be grown in other geographic regions safely.
Summary Date: September 1997
Objective: Conduct research and transfer technology to industry for use of genetically engineered trees in short-rotation plantation culture. The TGERC program is both adaptive and environmental in its orientation. Current research projects include: improving transformation efficiency; engineering herbicide resistance, insect resistance, and reproductive sterility; and assessing the genetic risks of transgenic poplars. These practical applications of genetic engineering can provide numerous benefits to biomass producers by: 1) enhancing tree growth, 2) allowing use of marginal land, 3) improving product quality, 4) decreasing management costs, and 5) reducing environmental impacts.
Approach/Background: Interspecific hybridization has led to the development of very fast-growing lines of hybrid poplar. Some of these hybrid lines are now being grown in large commercial plantations throughout the U.S. and Canada. Poplars are primarily grown for pulp and biomass, but are also valuable for removal of agrochemicals from groundwater, bioremediation of polluted sites, and for stream bank stabilization and restoration in riparian buffer strips.
Genetic engineering will improve poplar’s utility for short-rotation intensive culture. Although hybrid poplars have tremendous growth potential, they are very susceptible to insect herbivory, vegetative competition, and a variety of pathogens. Genes are now available to ameliorate these shortcomings; introducing them into commercially important hybrid clones will greatly enhance their usefulness.
Poplar is an ideal model system for woody plant genetic manipulation because it is easily transformable and regenerable, and has a small genome. Several poplar clones are readily transformed with Agrobacterium tumefaciens. Techniques recently developed in our labs have greatly improved the efficiency with which previously recalcitrant, but commercially important, clones can now be transformed and regenerated.
Status/Accomplishments: Using our improved Agrobacterium-mediated transformation protocol, coupled to an indirect organogenic regeneration system, we have achieved transformation efficiencies of up to 10% in a variety of recalcitrant hybrid cottonwood clones. We have also shown that matrix attachment regions (MARs) enhance and stabilize transgene expression. These elements should eventually improve transformation efficiency by increasing selectable marker gene expression. We are in the process of testing a copper-inducible gene expression system in hybrid poplar. This will allow for transient expression of genes that may improve transformation efficiency further by increasing a cell’s competence to differentiate.
Previously we generated 79 lines of transgenic aspens and cottonwoods that are resistant to glyphosate, the active ingredient in the herbicide Roundup®. These transformants contain two glyphosate resistance genes, and have been studied at three field sites over the past two years. A majority of these lines has demonstrated high levels of resistance and no detectable growth loss after multiple Roundup® applications. The three transgenic lines with the fastest growth and highest resistance to Roundup® are now being grown in stool beds to produce sticks for a long-term management study to be initiated in the spring of 1998. In addition, we are testing individual glyphosate-resistance genes in a single clone of cottonwood for their ability to impart resistance and for use as a selectable marker. We have also begun field-testing hybrid aspens that have been engineered for resistance to glufosinate, the active ingredient in the herbicides Finale® and Liberty®.
Recently we developed 77 independent lines of poplar (67 cottonwood, 10 aspen) that contain the CRY3A gene from Bacillus thuringiensis. The T-DNA of the genetic construct used to produce these transgenics was bordered by MARs. Nearly all of the lines tested show strong resistance to the cottonwood leaf beetle in laboratory assays. After screening the remainder of our transgenic lines, we will establish an insect-resistance field trial in an area with a large resident population of insects.
We believe that engineering reproductively sterile trees will facilitate regulatory approval and public acceptance of transgenic plantations. In addition to the several hundred transgenic lines previously developed, we have recently begun field-testing AP1-DTA transgenics. To date, we have cloned four floral homeotic genes from black cottonwood. Genomic clones for two of the most recently cloned genes, PTAG1 and PTAG2, have now been sequenced. In situ hybridization shows that these genes are expressed in floral meristems prior to the differentiation of floral organs. We intend to incorporate these poplar genes into our sterility program soon.
When floral promoters are used to engineer sterility, the effectiveness of those systems cannot be evaluated until transgenic plants flower. Therefore, conventional methods and floral homeotic genes are being studied for induction of early flowering. One construct containing 35S-LEAFY has caused flowering within months of transformation in both cottonwood and aspen. This will greatly speed studies of floral sterility.
The extent of gene flow from hybrid plantations to the wild is being studied. The information gathered will be used to develop a model to predict the impacts of deploying transgenic trees. Results of controlled crosses, conducted in cooperation with the Poplar Molecular Genetics Cooperative, reveal that the triploid clones we are genetically engineering have a high level of innate sterility. These same triploid clones are also less competitive than their wild relatives. Finally, we have identified several RAPD primers that amplify products which will allow us to conduct paternity analyses on hybrid seedlings.
Summary Date: September 1997
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File Created: October
13, 1997; Last updated: Thursday, 11-Nov-1999 10:23:34 EST