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W2168: Environmental and Genetic Determinants of Seed Quality and Performance

Annual/Termination Reports (SAES-422): [02/06/2009] [08/19/2010] [08/09/2011] [05/01/2012] [06/11/2013]

Date of Annual Report: 02/06/2009

Report Information:
  • Annual Meeting Dates: 01/16/09 to 01/17/09
  • Period the Report Covers: 10/2007 to 09/2008

  • Participants:
    Brief Summary of Minutes of Annual Meeting:
    Meeting opened by Chair: Hiro Nonogaki Oregon State University.

    Introduced Andy LaVigne: President and CEO of the American Seed Trade Association.

    Hiro Nonogaki asked for approval of the 2007 W-1168 Annual Meeting Minutes Kent Bradford: moved and Miller McDonald seconded. Minutes passed unanimously. Bradford and Welbaum will be on the committee for determining executive officers.

    The site selection committee will be Alan Taylor and Bob Geneve.

    Call for proposals of discussions for further meetings like the translational seed biology meeting.

    Discussions: Florida agriculture, ASTA, seed research, graduate education, collaborations (industry/government/universities and physiologists/breeders/geneticists), regional seed production industries, specialty groups, general education, distance learning, grants (World Bank, Gates, National Needs USDA, NSF), seed biology recognition, seed health & performance, seed health & conservation, germplasm, crops, identity & visibility, research focus, plant breeding.

    State reports (CA, MI, FL, OH, NY, TX, KY, SD, VA, OR).

    Detailed narratives for outputs and outcomes/impacts.

    Detailed paragraphs are included in the minutes attachment.

    A series of genetic experiments reached the conclusion that the rice qSD12 QTL controls seed dormancy through the embryo or endosperm tissues.

    Four wheat populations of about 700 double haploid or recombinant inbred lines were evaluated for seed dormancy and resistance to pre-harvest sprouting.

    Two QTL for germination heat sensitivity were mapped on the short arm of chromosome 3A (QGhs.osu-3A) and the long arm of chromosome 4A (QGhs.osu-4A) in Intrada x Cimarron RIL population of winter wheat. These two QTL were also mapped in Jagger x 2174 RIL population of winter wheat in similar genomic regions.

    The maize RILs (IBM 302) exhibit poor field emergence. Laboratory studies were conducted to find QTLs associated with seed vigor. The outputs from these studies are QTL maps from germination under low temperatures. Analyses of QTLs across a range of biologically important germination temperatures will be conducted (IA).

    Soybean seed with low raffinose, stachyose, and phytin are desired for feeding non-ruminant animals, but may have reduced agronomic quality. Research results provided information on the composition of low-raffinose, low-stachyose soybean seeds with normal phytin composition.

    Previously a major quantitative trait loci (QTL) termed Htg6.1 was identified that confers the ability for lettuce (Lactuca sativa L.) seeds to germinate at high temperatures (up to 37°C) in an accession of L. serriola (UC96US23) in a recombinant inbred line population derived from a cross with L. sativa cv. Salinas. Multiple near-isogenic lines (NILs) were developed in the Salinas background carrying an introgression containing the QTL locus. Multiple NIL families were utilized to demonstrate that this genomic region conferred increased upper temperature limits for germination. Additional fine mapping of the QTL locus using single feature polymorphism (SFP) markers developed using a lettuce microarray identified a gene encoding an enzyme in the abscisic acid (ABA) biosynthetic pathway (LsNCED4) that localized to the center of the QTL interval. Expression analyses were conducted on over 80 genes in the abscisic acid (ABA), gibberellin (GA), ethylene and light regulation pathways in relation to thermoinhibition in different genotypes. Gene expression studies have clarified some of the regulatory interactions among ABA, GA and ethylene in regulating seed responses to light and temperature (CA).

    Modifying the maternal plant environment has significant effects on lettuce seed quality. By controlling specific environmental conditions during seed production of the commercial lettuce cultivar Tango it was possible to modify seed size, germinability, thermoinhibition, photodormancy and storability. In these studies, one of the first questions to be addressed is if the effects of light quality on storability are observed in other lettuce genotypes and, more importantly, in other species. Additionally, finding a significant correlation between light spectrum quality of the maternal environment and storability in a wide range of species would be important for understanding the seed bank dynamics of native and weed plants. Basically, it would mean that seeds produced under a canopy shadow (i.e. lower R:FR ratio) have higher storability and are better prepared to remain viable for longer periods of time (OH).

    Early events in sugar beet germination were investigated using quantitative PCR. One hundred ninety two gene products were tested for expression at 0, 24, 72, and 96 hours post-imbibition. Differentially expressed genes generally showed expected patterns of expression, however results are preliminary (MI).

    Seed germination as well as longevity in dry storage are attributes affected by the testa (seed coat). The brownseed1 (bs1) gene in tomato is unknown. Recombinants between Solanum esculentum and S. pimpinellifolium were subjected to PCR with a variety of primers generating CAPS, dCAPS, and SSLP markers to continue fine mapping bs1 (KY).

    Germination of orchid seeds is more successful when seeds are harvested from immature pods. Protocols for decontaminating orchid seeds with calcium hypochlorite for propagation through tissue culture have been optimized. Orchid seeds can survive freezing in liquid nitrogen and therefore can be stored in genebanks cryogenically. In collaboration with Dr. Zhiwu Li, orchid was genetically transformed with a hairy root expression cassette and this work is continuing to genetically transform orchids for physiological studies. The orchid caraspace is a thin barrier to macromolecule diffusion and it may control orchid seed germination (VA).

    To identify DNA sites to which AGL15 binds in vivo, a chromatin immunoprecipitation (ChIP) approach was used to immunoprecipitate AGL15 and associated DNA fragments. In a ChIP-on-chip approach, the DNA recovered from immunoprecipitation using AGL15-specific antiserum or preimmune serum was converted to probe to hybridize to the Affymetrix GeneChip® Arabidopsis Tiling 1.0R Array. This allowed nearly global mapping of in vivo binding sites for AGL15 that numbered ~2000. Affymetrix ATH1 arrays were used to investigate gene expression changes in response to accumulation of AGL15/18. We are currently analyzing results to identify genes that may be directly regulated by AGL15 and those which may be farther downstream in the regulatory network.

    The roles of three unique Tandem Zinc Finger (TZF) genes in seed development and environmental responses in Arabidopsis thaliana were studied. Several sugar responsive TZF genes were identified using microarray analyses. Sequence analysis has revealed a family of 11 genes in the Arabidiosis genome that contains this unique TZF. It is interesting to find out the roles of these genes because the molecular mechanisms underlying ARE-mediated mRNA decay are unknown in plants (OH).

    Arabidopsis seed mutants completing germination were characterized. Yeast-two hybrid, western blot, single- and double-mutant analysis have determined that the F-BOX protein COLD TEMPERATURE GERMINATING10 (CTG10) results in the destruction of PHYTOCHROME INTERACTING FACTOR1 (PIF1) (KY).

    DELLA proteins are negative regulators of GA responses including seed germination, stem elongation, and fertility. GA can stimulate GA responses by causing proteolysis of DELLA repressors by the ubiquitin-proteasome pathway. This destruction requires GA biosynthesis, three functionally redundant GA receptors GIBBERELLIN INSENSITIVE DWARF1 (GID1a, GID1b and GID1c), and the SLEEPY1 (SLY1) F-box subunit of an SCF E3 ubiquitin ligase. Using sly1 mutants in which DELLA proteins remain stable after GA application, we found that GA regulates DELLA repressor activity by a mechanism distinct from protein destruction. Overexpression of GID1 genes rescued the dwarf and infertility phenotypes of the sly1 mutants without altering accumulation of DELLA proteins RGA and GAI. This rescue required GA biosynthesis and the presence of a functional DELLA motif in RGA and GAI. Both the DELLA motif and GA are required for the protein interaction of DELLA protein and GID1. The sly1 mutants display a less severe dwarf phenotype than the GA biosynthesis mutant ga1-3 or the gid1a gid1b gid1c triple mutant despite the fact that sly1 mutants accumulate far higher levels of DELLA protein. Based on double mutant analysis, it appears that both intermediate phenotype and high level DELLA accumulation in sly1 mutants require GA and a functional DELLA motif. These results suggest GA-bound GID1 can block DELLA repressor activity by direct protein-protein interaction with the DELLA domain, and this interaction may lead to increased DELLA accumulation (WA).

    To identify and characterize key transcription factors potentially associated with seed germination and stand establishment, miRNA regulated genes were characterized. Results suggest that miRNAs play critical roles in many different stages of plant development including seed germination and seedling growth.

    Objective 2. Determine and model the biotic and abiotic factors affecting seed germination, seedling emergence, and establishment of sustainable populations in natural and agro-ecological systems.

    Sensitivity of mature soybean seeds to imbibitional chilling were determined for low & normal raffinose and stachyose seeds, low raffinose, stachyose, and phytin seeds. Research results provided information on the sensitivity of low-raffinose, low-stachyose, low-phytin soybean seeds to imbibitional chilling injury (NY).

    A population-based threshold model was applied to quantify the responses of barley seed germination to oxygen availability, ABA and GA. The ability to quantify the sensitivity of seed germination to combinations of oxygen, ABA and GA revealed that oxygen availability and hormones interact to regulate germination, particularly in intact cereal grains (CA).

    Pritchardia remota (Arecaceae) is an endangered palm endemic to Hawaii. Management plans call for ex situ conservation of this species. However, the ability to store seed using conventional methods has not been investigated. Research on palm seed conservation was performed. Embryos seem sensitive to stresses imposed by high levels of drying and do not exhibit extreme desiccation tolerance required for conservation in genebanks. Therefore, until the effects of enforced desiccation on potential embryo dormancy can be elucidated further the feasibility of storage in conventional genebanks remains questionable. It is concluded that P. remota embryos possess a storage physiology intermediate to the recalcitrant and orthodox types (FL).

    Summer farewell (Dalea pinnata) is a legume found in upland ecosystems throughout the southeastern United States and often used in seed mixes for restoration purposes. It also represents an opportunity for the emerging native seed industry in this region. Yet, germination characteristics for summer farewell are not reported. Several key outcomes from studies described below are evident. First, restoration practitioners should expect low initial germination for summer farewell after seed shedding or sowing of non-treated seeds. Second, low germination does not necessarily equate to failure since seeds of summer farewell have the ability to form a soil seed bank. Third, the majority of the seed population possesses physical dormancy but this dormancy can be readily broken through scarification. Fourth, the seed or seedling producer may increase germination by scarifying seeds. Finally, further studies dealing with seed development, polymorphism, and the role of fire on dormancy break are required to determine how these factors influence germination and recruitment of summer farewell (FL).

    The principles of PCR and Density Gradient Gel Electrophoresis have been applied to studying microbial seed ecology. This technique along with 16S gene sequencing will allow use to genetically profile microbial populations residing on seeds (VA).

    Objective 3. Develop, evaluate, and transfer technologies to assess and improve seed and seedling quality, health, performance, utilization, and preservation.

    Many seed crops have been identified as suitable for biofuel production producing a need for high quality seed lots that will produce optimum plant stands with the potential for maximum biomass production. Seed dormancy, suboptimal soil temperatures, and pathogens often lower seed quality and can negatively impact stand establishment. Seed testing methods provide an overall assessment of the quality of a seed lot; however, variants in the germination test method can have a major influence over final germination rates and percentages. Knowledge of the seed biology of species used for biofuel production is needed to understand factors or conditions limiting germination. Specific technology methods can be adapted or developed to optimize seed quality and stand establishment (NY).

    Because of its wide geographic distribution and high yield potential, switchgrass is considered a leading candidate as bioenergy crop. Poor stand establishment is a frequently encountered problem in growing switchgrass. This is mainly associated with high degree of dormancy of switchgrass seeds. Global gene expression analysis using rice long-oligo arrays were conducted to identify genes that showed differential expression in dry and germinating switchgrass seeds (TN).

    Sweet corn (Zea mays L.) seed has naturally low physiological quality compared to field corn. The use of priming treatments has been recommended to decrease the time between sowing and seedling emergence. In sweet corn seeds, hydropriming and osmopriming methods increased the uniformity of seedling emergence and reduced the range of days for germination. Priming evaluation is normally conducted by germination assays; however for vigor analysis of seed lots, more precise, quick and efficient tests are desirable (OH)

    Results obtained for sweet corn sh2 'SWB 551' and 'Obsession' hybrids demonstrated that high quality sweet corn seed lots respond positively to priming; however, priming is not beneficial for seed lots with medium physiological quality. The best treatment using the drum priming system was 36 h priming and for this duration, SVIS (vigor index ratio of 70 growth + 30 uniformity) produced similar results to seedling emergence assessments documenting this as a rapid seed vigor test that can identify the efficacy of priming treatments (OH).

    Proteomic analysis of the wheat grain has advanced our understanding of endosperm and amyloplast proteins. The gluten proteins, a complex collection of highly repetitive storage proteins, are of major interest since they determine the characteristics of extensibility, elasticity and gas-holding capacity that are unique to wheat flour doughs. Proteomics has provided new information on the identification and regulation of major metabolic pathways functional in the endosperm and amyloplast, knowledge that is fundamental to the understanding of both the quality and productivity of the wheat grain (CA).

    Seed proteins of sorghum are less digestible than those of other cereals and digestibility is exacerbated by wet cooking the meal or flour, which results in significant nutritional losses. To address this problem, the properties of two sorghum lines that have a common pedigree but differ in digestibility were analyzed. Consistent with results based on a ruminal fluid assay, the protein and starch of one line (KS48) was more thoroughly digested than that of the other (KS51) using in vitro assays based on pepsin and a-amylase. The indigestibility of KS51 relative to KS48 was shown to be due to (i) a greater abundance of disulfide-bonded proteins; (ii) presence in KS51 of non-waxy starch and the accompanying granule-bound starch synthase; and (iii) the differing nature of the protein matrix and its interaction with starch. The current findings suggest that each of these factors should be considered in efforts to enhance the nutritional value of sorghum grain (CA).

    Seed priming (controlled hydration followed by drying) is used to overcome thermoinhibition in lettuce and to speed germination in a number of species. Seed priming (prehydration and drying) is widely used in high-value seeds to improve seed germination uniformity and speed and to overcome dormancy, particularly in lettuce, where it increases the upper temperature limit for germination. Transfer of the trait to cultivars may have the added benefit of making them more responsive to priming treatments. (CA).

    Effects of aqueous slurry seed coatings were examined.

    Field and laboratory bioassays were conducted in multiple States (Delaware, Maryland, New York, Ohio and Virginia) to assess efficacy of thiamethoxam, and clothianidin + imidacloprid seed treatments and compare with efficacy of in-furrow applications of thiamethoxam, and imidacloprid.

    A growth chamber study investigated abiotic stress tolerance of artichoke transplants.

    A method to extract true seeds from beet fruits (seedballs) was enhanced. The method involves a brief soak in water that resulted in making the seedball pliable, deformation of the wetted seedball with firm pressure until the cap separated from the seedball, and teasing the true seed from the fruit. This method is being used to investigate mechanisms of seedling vigor and assessing seed quality in sugar beet during the critical imbibition, biochemical reactivation, and early growth of sugar beet seedlings (MI).

    A new instrument to measure respiration (oxygen consumption) of individual seeds in a 96-well format (Q2, www.astecglobal.net) was evaluated as a seed vigor test. Seed lots representing diverse quality levels and enhancement treatments were assessed in the instrument. Indices associated with the quality of seed lots were calculated based upon the parameters of the oxygen uptake patterns of individual seeds. The relationships of individual seed respiratory patterns were evaluated with respect to other indices of seed quality. While some technical improvements are still required, the Q2 instrument provides a useful tool for evaluating seed quality (CA).

    Vigorous and healthy seeds are an essential start toward successful crop production. Seed sanitation methods (disinfectants, heat treatments, biological protectants) may be more or less successful depending on factors such as seed vigor, seed maturity and initial levels of seed pathogens. Studies were conducted to evaluate (i) the effectiveness of seed disinfectants and chemical treatment in eliminating Clavibacter michiganensis subsp. michiganensis (Cmm), the causative agent of bacterial canker, on and in tomato seed, (ii) the effectiveness of seed treatment combined with biocontrol bacteria in eliminating Cmm on and in tomato seed, (iii) the influence of seed treatment on seed vigor and disease control effect in greenhouse tomato seedlings, (iv) the influence of seed treatment on seed vigor in storage, and (v) the influence of seed treatment on seed vigor (OH).

    The Seed Vigor Imaging System (SVIS) is a vigor test that evaluates seed performance by scanned images of young (three-day-old) seedlings evaluated by computer software. In SVIS, seeds are germinated at 25oC, the resulting seedlings scanned and their length and uniformity analyzed using software that computes an overall vigor index. This test provides a rapid and objective measurement of seed quality, and the images and vigor indices are stored and a data base developed for future reference. The objective of this study was to analyze the effects of priming treatment on sweet corn seeds using SVIS (OH).

    Seed technology and production training DVDs have been completed. Seed testing (importance of seed testing; seed quality; tetrazolium tests; genetic purity) and seed production modules (coffee/tropical forage grasses; sunflower; maize) are currently being marketed by the Society of Commercial Seed Technologists. (OH).

    A one-dimensional temperature programmable thermogradient table was designed with LED grow lights to utilize new technologies for seed testing. The case and lids are made extensively from polymers that reduce weight and are not susceptible to water damage like wood framed tables. The TASCO gradient tables expand capabilities for seed testing and research because temperature gradients can be programmed and seedlings can be grown using the LED grow lights. The LED lights provide wavelengths in the photosynthetic spectrum and do not burn out like conventional lights. Since water resistant polymers are extensively used in construction, the case is water resistance (VA).

    A protocol to rapidly view high resolution confocal images of purpletop (Tridens flavus) embryos in dormant and non-dormant (scarified) caryopses was developed. Decreased days to 10%, 50% and 90% of final germination percentage and increased length of radicle and widths of main axis and radicle were concomitant with rapid subsequent germination of non-dormant carypopses, but not of dormant ones. Thus, embryo observation using confocal microscopy may be a useful technique for establishing seed vigor (DE).

    Incubating seeds (florets) of Aristida stricta in glycerol, lactic acid, glycerol + lactic acid, or lactophenol for up to 24 hr failed to clear the embryo coverings enough to view TZ staining patterns. Use of a press test only accurately predicted that a seed was nonviable. Based on preliminary results, viability and germination of a seed lot can be accurately determined by conducting a press test, subjecting only filled seeds to a germination test (14 days @ 15/25oC in dark), and then conducting a press test on nongerminated seeds (FL).

    Priming Coreopsis floridana seeds in polyethylene glycol (PEG) +BA or SMP (emathlite clay)+BA resulted in 100% germination at 20 or 30oC in the dark. Germination of C. lanceolata at supraoptimal temperatures was improved by priming seeds in PEG+BA (FL).

    Impact Statements:
    1. A large number of genes that showed differential expression in dry and germinating switchgrass seeds were identified. Some of these genes may be critical for regulating dormancy and germination of switchgrass seeds. Further detailed functional characterization of these genes may lead to novel insight into the molecular mechanisms of seed dormancy and dormancy breaking in switchgrass. Such information may provide important knowledge and tools for genetic improvement of switchgrass as a bioenergy crop.
    2. The rice qSD12 locus is the first seed dormancy QTL that has been genetically characterized for seed tissue expression. The partial genetic map for the rice qSD1 region facilitates map-based cloning the gene underlying the dormancy QTL.
    3. Wheat seed quality research has provided a foundation both for plant breeders to search for specific traits and cereal chemists to seek a solution for dough quality. These are longstanding problems that have faced agriculture for decade.
    4. The phenotypic data obtained from the wheat seed dormancy segregating populations will be used to map the QTLs for resistance to pre-harvest sprouting.
    5. Soybean seed research results led to a new idea for improvement of field emergence of low-phytin soybean seeds. Increasing the concentration of total a-galactosides of myo-inositol, D-pinitol, and/or D-chiro-inositol may improve field performance of low-phytin soybean lines. Since low-phytin soybean has low galactinol concentrations (the galactosyl donor for galactopinitols and the raffinose family oligosaccharides), it is specifically proposed that an increase in fagopyritols, a-galactosides of D-chiro-inositol, may result in improved field performance of low-phytin soybean.
    6. The detrimental effects of freezing temperatures at the late stages of seed development and maturation can have a dramatic effect on corn seed quality. Freezing injury in corn causes irreversible physiological damage. Every four or five years, an early fall frost in Iowa causes substantial monetary losses for the seed industry due to lower seed germination and vigor. Yet, damaged seed does not show adverse symptoms immediately after harvest. Several months later, seed vigor will decrease below the market standards for the seed trade. Seed vigor is the expression of an aggregate of traits. QTL analysis is therefore a logical starting point to understand the genetics and biochemistry of seed vigor. The analysis of QTLs across a range of maize seed germination temperatures will explain some elements of the variation inherent in QTL studies.
    7. Grain sorghum (Sorghum bicolor L. Moench) ranks fifth in worldwide production among cereal crops, after wheat, rice, corn and barley. The U.S. is the number two producer and the number one exporter of sorghum, primarily to Mexico for use as animal feed. Popularity of sorghum is due in part to its ability to produce reasonable yields in warmer, drier regions. Because subsistence farmers in Africa and Asia cultivate sorghum widely as a staple food for home consumption, the crop is a principal source of energy and protein for millions of the worlds poorest. The research on sorghum seed quality has huge impacts on U.S. and global agriculture.
    8. In lettuce, most of the traits improved through domestication are associated with the leaf characteristics, while the seeds exhibit traits associated with wild plants, such as dormancy. A gene (LsNCED4) that is likely to be the basis of the trait has been identified and revealed some of the hormonal mechanisms that regulate germination and could be applied to control weeds or enhance crop emergence. Developing varieties whose seeds respond more strongly to priming will enhance seed performance under stressful field conditions.
    9. The advances in knowledge of how light, temperature and water availability affects lettuce seed vigor improved seed quality. Seed companies and growers will refine and adjust production methods to reliably produce lettuce seed lots of high vigor.
    10. Understanding seedling morphological and physiological responses during germination, growth and establishment under extreme temperature and drought conditions is important to develop seedling conditioning treatments to mitigate those stresses. ABA alone or in combination with other plant growth regulator (PGR) have the potential to improve transplant stress tolerance by regulating transpiration, enhance plant water status and by controlling transplant development.
    11. An in-depth understanding of gene expression during the critical early events of sugar beet seed germination is needed in order to identify key regulatory steps in seedling vigor, and identify mechanisms of seedling vigor under different germination regimes and in different genetic backgrounds.
    12. The new seed extraction method from beet fruits to allows access to the previously inaccessible true seed, which has been an impediment to assessing seed quality and biochemical events underpinning seed vigor in sugar beet.
    13. Seed respiratory rates during germination may provide rapid and simple indices to assess vigor differences among lots.
    14. The systemic insecticide seed treatments, thiamethoxam, and a combination of clothianidin + imidacloprid reduced cucumber beetle damage in cucumber and pumpkin. These insecticides were not injurious to seed. Thiamethoxam was labeled by Syngenta in 2008, and a label is being pursued for combination of clothianidin + imidacloprid by Bayer CropSciences with IR-4 and other regulatory agencies.
    15. The Seed Vigor Imaging System (SVIS) has provided a rapid and objective measurement of seed quality.
    16. In cooperation with TASCO LLC of Christiansburg, VA programmable one-dimensional thermogradient tables with LED lighting are being produced commercially for scientists and the seed industry (VA). The TASCO tables sell for significantly less money than other thermogradient tables that are commercially available.
    17. Dormancy classification studies in this project are helping to fill significant gaps in our knowledge of the world biogeography of seed dormancy. The research on Convolvuloideae is especially the first time for the phylogeny of seed dormancy in a large complex subfamily being elucidated.
    18. Understanding the physical and physiological mechanisms that control seed dormancy can provide approaches to modify it for agricultural applications. In barley, for example, uniform germination is critical for the malting process. Knowledge of the roles of seed covering tissues and hormones in regulating dormancy and germination will reveal new methods to attain uniform germination without reducing dormancy to the point that pre-harvest sprouting is problematic.
    19. In terms of global economic importance the palm family, Arecaceae, is categorized with Fabaceae and Poaceae. Palms are widely distributed primarily throughout sub-tropical and tropical ecosystems where they are highly utilized by indigenous people, present sustenance for many animals, and provide vital ecological functions. Despite these benefits, the genetic diversity of many palm species is threatened due to habitat loss and overexploitation. Furthermore, ex situ conservation of Arecaceae within gene banks is hampered due to a poor understanding of palm seed biology. The palm research in this project has provided useful information for palm seed conservation.
    20. Native wild flower seed research will enhance the establishment of sustainable roadside populations in a cost-effective manner. The presence of roadside populations of native wildflower ecotypes will reduce mowing costs when mowing cycles are timed to promote reseeding.
    21. The press test in combination with a germination test to determine viability and germination of Aristida stricta seeds is a protocol that may be more cost effective than the tetrazolium (TZ) test in combination with germination testing.
    22. Priming seeds of Coreopsis floridana and C. lanceolata provides a potential means of enhancing germination and establishment under field conditions, even under high summer temperatures.
    23. Seed lot quality has surfaced as a key issue in commercially available gamagrass seed. Significant improvement in germination and stand could result from density separation for heavier seeds. In addition, H2O2 could be an effective alternative to stratification with the advantage being a reduction in the time required for dormancy release (i.e. 12 hr for H2O2 compared to 4 to 6 weeks stratification.
    24. The orchid seed research influenced the research of other orchid researchers who used to view the caraspace as an insignificant tissue that simply got in the way of orchid embryo research. Transformation research on orchid, once thought impossible, it now becoming more routine. Calcium chlororide is being adopted to sanitize orchid seeds for propagation. Other research groups are exploring cryopreservation of orchid seeds and the role of the caraspace in controlling dormancy/germination.
    25. The study of microbial seed ecology using the principles of PCR and Density Gradient Gel Electrophoresis is anticipated to provide valuable information on seed transmitted organism because both culturable and nonculturable organisms can be detected.
    26. A means of regeneration, either by organogenesis or somatic embryogenesis, is necessary for genetic engineering for most plants. However, somatic embryogenesis is poorly understood. Somatic embryogenesis serves as a model for zygotic processes that are also largely a mystery and difficult to study because early embryo development is relatively inaccessible. A better understanding of developmental mechanisms controlling somatic embryogenesis including AGL15 will aid regeneration of recalcitrant species in culture and contribute to an understanding of seed development.
    27. A better understanding of the stresses imposed upon the orthodox seed proteome may provide techniques to better handle seeds during late development so as to prepare them to rapidly and efficiently repair this damage.
    28. Identification of TZF genes in Arabidopsis advanced our knowledge on the development and expression of seed quality.
    29. The research on the DELLA seed germination repressor proteins has determined that GA receptor GID1 can inactivate the DELLA repressor RGA in the absence of protein destruction and provided new insight into hormonal regulation of seed germination.
    30. Characterization of miRNAs in seeds has provided new insights into the mechanisms of gene regulation during seed development and germinaiton. miRNAs are conserved among wide ranges of plant species. Therefore, the outcomes from Arabidopsis seed miRNA research can be applied to crop species.
    Last Modified: 20-Mar-2009

    Date of Annual Report: 08/19/2010

    Report Information:
  • Annual Meeting Dates: 07/09/10 to 07/10/10
  • Period the Report Covers: 10/2009 to 09/2010

  • Participants:
    Brief Summary of Minutes of Annual Meeting:
    Minutes of the ASRA (W-2168) Annual Meeting July 8-11, 2010:

    8:35 am call to order by Bruce Downie followed by introductions of participants. Downie continued with statement to members outlining accomplishments that included: 1) selecting a name for W-2168 group that would not change with every re-write of the project, American Seed Research Alliance was the selected name; 2) setting up a group webpage; and 3) submitting an NSF Research Coordination grant. The grant was not funded. This highlights need to obtain other sources of funding that include collaborations with other groups. Also, try to liaise with ASTA to leverage federal funds. After the statement Downie sought approval of minutes; 1st and 2nd motions for approval of minutes by Bradford and Bennett, respectively.

    Mark Bennett and Hector Pérez were selected as new President and Vice-President, respectively. Kent Bradford and Marc Cohn were appointed to the Secretary Nomination committee. Downie re-iterated need to email state reports, using SAES-422 format, and minutes to him.

    Taylor, Bennett, and Pérez were appointed to Host Site Selection committee.

    Downie opened the floor to discussion of special topics from membership. Bradford requested time on Saturday to discuss ASTA request for input on topics to present to Roger Beachy (USDA-AFRI). Bradford suggested an ASRA summit sponsored by ASTA and creation of a bulleted list of seed-related items. General consensus among members that ASRA should have some input and provide feedback on agenda related to seed research at federal level.

    Cohn described challenges in building collaborations due to diversity of mandates and interests among members. Moving forward a unifying theme could be the need to perpetuate the species of seed scientists since general lack of awareness of people who know seeds as propagules. Focusing on seeds as propagules allows for wide interests.

    Bishnoi suggested seeds are integral to a diversity of new research areas from crop biofuels to medicinal plants and these may be areas to focus on. Moreover, it is important to include minorities in collaborations. 1890 schools are rich in minorities and faculty with interests in seed biology and should be considered for partnerships.

    9am Welcome and Introductory remarks from Doug Buhler MSU Introductory remarks focused on: need to define role/expectations of ASRA, finding commonality and building collaborative research projects (e.g., NCDC has committee specifically to write grants; strength in the multi-state system), name of group can be considered as a brand, focus on asking what can we do specifically to match our needs.

    Buhlers presentation focused on state of MSU Ag experiment station and opportunities.

    Questions from the members revolved around how to keep seed science alive in the face of mergers and budget cuts, issues with distribution of appointments among units, and replacement of Dr. Copelands position and diversion of resources to teach more about seeds.

    Buhler  Dr. Copelands position will not be refilled. Positions are in danger of budget axe, but opportunities exist to link funding with contemporary issues. Look at cluster hires and find out if seed science can participate. Major problem revolves around not finding qualified people to fill positions. Need to hire the best then give them time to develop. Innovation is required. Reorganization can present more challenges than opportunities. There are costs associated with mergers that can take a decade to smooth out (e.g., lost grants and publications). MSU tries to be strategic with distribution of appointments. Administration views experiment station as a major piece of the university. However, big budgetary changes make deans nervous.

    As voted by members, floor opened for discussion of special topics for 1 hr. Leskovar reported upcoming ISHS meeting with a session emphasizing seeds transplants and stand establishment to occur in Brazil in 2012; Welbaum announced seminar/symposia on relationships between seeds and seed microbes planned for ASHS meeting in Portugal on Aug, 20-26th.

    Focus of discussion was on potential funding sources for ASRA. A recurring theme was the need for seed education at all levels. Pérez suggested looking in NSF GK-12 funding mechanism as this has the dual purpose of grad student funding and potential for seed biology education projects; Downie suggested USDA-National Needs program as another potential funding mechanism, grad students receive training by seed biology faculty; Bradford mentioned project in the wheat community that trains cohorts at different institutions, each institution interacts, perhaps this could serve as a model for ASRA funding efforts, emphasized need to make a case on importance of seeds to those that make priorities; Welbaum discussed published review on distance learning in seed biology found in Seed Technology, Agronomy Newsletter and New Seeds, need to consider integrating distance learning in education projects, but this can be affected by retirements.

    Bradford/Downie/Cohn/Geneve/Bisnoi discussed creation of a center of excellence in seeds, further discussion would be carried out during break. Cohn emphasized need to focus on fundamentals of seed biology then the flavor du jour can be added (e.g., biofuels) to market an agenda. Concern was expressed about departments only funding in high priority areas without interaction of seed person and tenure issues with difficulty in obtaining funds, this highlights the need for collaboration. An international component would strengthen the project.

    Geneve/Taylor/Downie discussed opportunities and challenges with the USDA-SCRI funding mechanism and questioned whether it would be the right mechanisms for such a diverse group. Pérez questioned if it would make sense to have Feds consider seeds as a specialty crop, perhaps including the suggestion as one important bullet point to ASTA and AFRI. Lemaux/Bradford discussed efforts on submitting a Seed CAP grant, challenges associated with CAP grants revolve around focus on one crop, this funding mechanism must include a research, education, and outreach component, perhaps need to stay clear of controversial topics, USDA concerned with how much has been invested with few current examples.

    Taylor/Geneve/Leskovar/Bradford shifted back to SCRI, mechanism sounds interesting, but foresee difficulty including members under same umbrella, industry buy-in is critical, a 50% match is required, huge amount of effort to develop a grant for this mechanism.

    Break 10:35am

    Resume 11:00am

    Downie/Rachael Naegele (McGrath grad student)/Geneve discussed National Ag in the Classroom as a means to increase seed biology awareness and as a possible addition to ASRA education grant opportunities, Ag in the Classroom is already established in every state, ready resources that can be made use of and improved, ASRA should identify Ag in the Classroom representatives in each state, provides good framework for collaboration.

    Bradford/Bishnoi/ Cohn discussed opportunities and current successes with USAID and HortCRSP projects, membership may consider these as funding mechanisms, need to target developing countries, major emphasis on poverty alleviation and African and South Asian countries, website for CRSP project is www.hortcrsp.ucdavis.edu.

    State Reports

    Dr. Peng  looking for collaboration opportunities, Epigenetic control rice seed grain filling how it affects nutrition; lab works on proteomics; willing to collaborate on protein identification; working on biomass crops to identify proteins seeking collaboration. Questions: Bradford- storage proteins more enriched, what stage of development? Reply: will still get enriched, not every cell highly expressed, very early stage of seeds about 6d. Cohn  this would be a good control, test later in development; Peng  starch accumulation later in development a challenge.

    Dr. Pérez  research program focused on seed development, dormancy and germination ecology; 2 peer-reviewed publications for 2009; 6 conference proceedings 3 with grad student as first author; 2 national presentations, 1 with grad student as lead author; 5 regional presentations 2 with grad student as lead; in 2010 3 publications 1 with grad student as lead author; expect another 3 in 2010; 2 conference proceedings, 1 international presentation and 2 regional presentations; seeking advice on starting seed technology program. Questions: Geneve has the role of fire been investigated in terms of dormancy relief? Reply No. Cohn  what are the colors of seeds at dispersal? Reply  We have identified 2 color morphs at dispersal

    12:10pm lunch; Resume 1:15pm

    Dr. Copeland  described book published in 2008 dealing with up-to-date taxonomy of seeds from numerous species; following up this effort with a seed ID website that will be hosted by MSU; website will contain color photos; author seeks photos and input. Also, working on new book titled Principles and Practices of Seed Testing. Downie/Lemaux/Bradford had questions regarding including a scale with photos, interest in SEM pictures and inclusion of dichotomous keys. Replies: including a ruler with image is fine, need to include magnification; welcome good quality SEM pictures; dozens of keys used in book.

    Dr. Cohn - exploring recalcitrant seed death with Spartina model systems; lipid oxidation spikes, antioxidant stays high until peak of lipid peroxidation, antioxidant pool being depleted, as antioxidant depleted increase in protein carbonylation, increases dramatically after CWC, not correct; what is the control for drying recalcitrant seeds? Do same rxns occur in orthodox seeds? DNA fragmentation does not occur as S. alterniflora is desiccated, but let them sit for time and DNA fragmentation occurs, may be post-mortem effect, little bit of moisture may facilitate fragmentation, do seeds die during drying or on rewetting? 2D gels 54 protein differences, hope to identify some putative gene products, molecular probe stains, a 24 kDa protein in dried S. pectinata (orthodox) not found in dried S. alterniflora, dormancy proteomics in Spartina: 61 protein spots differ during cold differentiation. No change in CWC in relation to drying temperature. Bisnoi/Pérez asked about independence biochemical reactions and seed death and potential lack of dry matter accumulation as factor in recalcitrance. Reply: increase in TBARS in orthodox and recalcitrant species can be an artifact; have not investigated dry matter accumulation.

    Dr. Bradford  compositae genome project (NSF funded), gone to 15k map locations, have several markers for lettuce, about half genes on map, main focus on new project to develop common inbred line of high temperature tolerance; Can do transciptome analysis, weighted correlation network analysis takes all data stores in modules that co-expressed and behave more or less similarly; Cohn - when discriminating filter is removed how do maps link change? e.g. from 5 to 10%; Bradford  it is sensitive to those limits; are proteins from both alleles functional? Knock outs of LsNCED4 tilling mutants; functional test for seed thermoinhibition with Salinas LsNCED4 promoter and coding region transformed to UC96US23; Astec Q2 machine continues to be improved; Improved seed storage (HortCRSP), zeolite a new form of desiccant, can go through repeated drying cycles and still be effective, make very simple seed storage system for low volume users in developing areas; Cohn  can it be scaled? Bradford  yes; Seed Biotechnology Center staff is expanding, Seed Biology Production and Quality, PBA Class II started 2nd branch in Europe, curriculum Delphi study to develop a curriculum in plant breeding what do plant breeders need to know today? What do they need to know for industry? SEED School educate new seed industry employees in a five day course; Bradford/Downie  Pepper chip has been developed, available for researchers; Gu  any ABA linkage? Bradford  hope to get there, trying to look at promoters perhaps one sensitive to temperature the other not to track back to temp signal. Cohn  in light of ABA receptor components have you gone back over data, Bradford  there are some homologues, have not done any real work with it. One component may be related to dormancy. Misra  suggestions for next step using wet and dry soybean seed as wet seed loses and dry seeds pick up moisture; Bradford  need to kill seeds, problem 80% RH 40C seeds cant dry, cant use ambient air to get dry seeds need to heat them, but would kill them; Misra  use dry grain as desiccant; Bradford  seed drying system needs to be closed system.

    Dr. Lemaux  Barley with improved malting quality, wheat with improved dough quality and reduced allerginity; thioredoxin as redox sensor, engineered barley to overexpress thioredoxin h in protein bodies of the endosperm, by overexpressing thioredoxin could increase pullulanse activity and germinate 1 d earlier; Chinese collaborators looked at consequences of silencing trxh to delay germination (pre-sprouting), most dramatic effect on pre-harvest sprouting, trxh linked to scarecrow promoter expression limited to a single cell layer in root, GFP moves throughout the root this particular thioredoxin is a communicator of redox status in seeds, Trxh9 is mobile suggests role in cell-to-cell communication. Cohn  is preharvest sprouting in one variety? Lemaux  found in 3 different Chinese varieties. Cohn to group  preharvest sprouting before maturation drying? Group no; with climate change more preharvest sprouting; Gu  how difficult to transform wheat; Lemaux  difficult to do, can be done with agrobacterium, less difficult with bombardment, dont want to get rid of all trx; Unidentified grad student  all stocks publically available? Lemaux  need to contact people in China, its difficult moving plant materials between countries especially GMOs, this works in the field; Bradford  problem with white wheat can grow it but too risky due to possible pre-harvest sprouting; Gu  preharvest sprouting in northern great plains; Misra  happens in seed corn too, not able to get funding due to intellectual property issues, how to get funding look at it for productive seed get male and female to germinate at same time; Downie  At trxh travelling, look at GFP in endosperm? Lemaux  no.

    Dr. Gu  progress on seed dormancy in rice and wheat, comparative genetics of seed dormancy between tropical and temperate ecotypes of weedy rice, divergence in seed dormancy between weedy rice and cultivars; during domestication 80% of dormancy genes eliminated, qtl clusters provide suggested coevolution/adaptation of weedy traits; map based cloning of qSD7-1affects pericarp color, high ABA content but decreases later in development, SD7-1 controls dormancy and pericarp color by switching on/off other gene traits, what else regulated by this gene, detected 5 genes involved in pericarp color, conserved seed dormancy function among structurally different alleles, Mapping seed dormancy QTLs in wheat, 11 putative seed dormancy QTLs detected from a DH population; Cohn  temporal processes come into play when seeds are dormant from the time they are viable 7-10d coincides with ABA peak, seeds are green, red coloration comes much later in development, dormancy there before red pigment is there; Bradford  loss of dormancy genes? Gu  pleiotropic effect one gene controls multiple functions; Unidentified grad student  red pigment controlled by single gene? Gu  natural variation controlled by single gene retain white allele; Grad student  3 red genes control color? Gu  wheat 3 pairs of genomes rice diploid; Misra  rice take lots of water to grow how to connect water use of rice? Look at rice that will germinate with less water or evapotransporative consumption. Gu  why weedy rice so popular in rice growing areas, red color type more successful to colonize areas. Cohn  is mutant SD7-1 w/ red pericarp display seed shattering? Gu  dormancy phenotype separate from shattering.

    Dr. Downie  Seed Biology Group at UK; Baskins had numerous publications in 2009 and will be presenting a seed ecology workshop in New Caledonia during 2010; Downie submitted NSF Research Coordination Grant, expanded membership of ASRA, developed ASRA website; Sharyn Perry had two publications and received two grants.

    Dr. Welbaum  IHC Science and Horticulture for People seminar molecular profiling of microbial communities associated with seeds (Aug 22-27th 2010), screen Cucurbit germplasm for HGR to effectors in Agrobacterium, bacterial pathogens that affect human health role on seeds, using DGGE for whole populations on seeds, some bacteria present on the seed get carried on leaves as plants develop in the field, can production area be correlated to bacterial species? Soil priming using disaccharide solution added different concentration of sugars, bacteria in soil that produce glucose based polymer together increases water holding capacity, wont have affect with sterilized soil. Cohn  have any other lots been tested? Welbaum  two lots worked 20 did not; Group response  need inoculate media; Taylor  set up to do taxonomy work? Welbaum  did sequencing have tons of data that need to be worked through, been limited by what can be cultured; Taylor  all relatively benign to seed? Welbaum  looking for antagonists that can inhibit other bacteria; Bennett  looking at sprouts? Welbaum  yes; Bennett  very little is known huge differences in species that would show up; Bradford  seed industry trying to avoid that E. coli can be promulgated by seeds; Bradford  a paper where they inoculated seeds very heavily and some could be found on young leaves. Misra  how are seed courses? Welbaum  students from other campuses taking seed classes, undergrad seed course about 15 students every spring

    Membership voted to reconvene at 8am on Saturday.

    Discussion by Taylor/Bennett and group on where/when to hold next meeting; larger membership should be polled, but this time seems to work for most; look to increase diversity of group (e.g. more 1890s) at next meeting. Bennett made motion to hold meeting same time next year, 2nd by Misra.

    Meeting adjourned 5:35pm.

    W-2168_ASRA annual meeting minutes -- Kellogg Center, Michigan State University, East Lansing, MI July 9-10th, 2010. SATURDAY In attendance: Zhachua Peng, Mississippi State University, Udai Bishnoi, Alabama A and M University, Peggy Lemeaux, Univ. California Berkeley, Daniel Leskovar, Texas A and M University, Bruce Downie, University of Kentucky, Marc Cohn, Louisiana State University Agricultural Center, Kent Bradford, Univ. California Davis, Greg Welbaum, Virginia Tech University, Mitch McGrath, Michigan State University-USDA, Xingyou Gu, South Dakota State University, Marc Bennett, The Ohio State University, Bob Geneve, University of Kentucky, Alan Taylor, Cornell Agricultural Experiment Station, Manjit Mishra, Iowa State University. Student: Yuanjie Su - MSU grad student with Janet Lewis, Crop and Soil Science. Yuanjie is interested in breeding wheat.

    Not present Saturday but present Friday: Hector Pérez, University of Florida, Guests: Larry Copeland and Doug Buhler Students: Rachel Naegele - MSU grad student with Mitch McGrath, MSU-USDA. Rachel is interested in seedling vigor. Prannda Sharma - Eastern Michigan University doing a summer internship with Mitch McGrath, MSU-USDA. Prannda is interested in cell walls.

    Reconvene Saturday morning at 8:15.

    Marc Bennett, Daniel Leskovar, Robert Geneve, Udai Bishnoi provided their state reports.

    10:10 Break

    10:40 reconvened with Mitch McGraths and Manjit Misras state reports. Broke for Lunch at 12:10.

    1:20: Reconvened.

    Nomination committee was pleased to announce that Xingyou Gu, South Dakota State accepted their nomination. His candidature was moved for acceptance by Bruce Downie and Seconded by Marc Cohn. The motion for Dr. Gu to move into the secretary position was put to the vote and passed unanimously. Congratulations Xingyou, our new Secretary.

    The Site committee reported next and Alabama Agricultural and Mechanical University, Normal, AL, was chosen as the potential site for next years meeting. This will hinge on Udai obtaining the permission and financial support necessary to host the meeting at Alabama. Bruce will work with Udai to develop a letter requesting permission to have the W-2168 meeting at Alabama A&M University next summer. Back-up plan is to have the meeting at Iowa State University with Manjit Misra hosting. The motion of having Alabama host the meeting was moved to be accepted by Marc Cohn and seconded by Alan Taylor. A vote was conducted which passed unanimously. Alabama next summer.

    A call for any new business brought forth a request to have the outline for the State Reports sent by Kathleen, resent by Bruce Downie to the membership.

    This also brought us to discussing strategy and tactics for obtaining funding from some/any source to forward the research/education/extension efforts of individuals or sub-groups of the membership. It was the consensus of the group that funding for the entire W-2168/ASRA would not be possible through any mechanism currently in place. So, we should focus on sub-groups within the membership and support their efforts as much as we can.

    The overwhelming opinion was that the membership must work closely with the American Seed Trade Association (ASTA) if our opinions are to be considered. Otherwise, the ASTA will move forward with its own agenda and we will be left on the sidelines, irrelevant.

    There may be something that could be done with NIFA/AFRI to identify centers of excellence for seed science but this is long-term. More immediately, working through ASTA, it might be possible to highlight some areas of seed research that could be supported.

    The Gates foundation is interested in securing the food supply (they paid to have the seeds flown to the Norwegian Svalbard Doomsday vault) and educating the African Nations on all aspects of agriculture that might work in these nations. As an assemblage of agricultural experts with seeds as a focus, we have a unique knowledge base from which to draw for Educational efforts. Both the Gates Foundation and the NSF-BREAD grants require boots-on-the-ground to implement agricultural research and innovation. To this end, Bruce was delegated to start a list of the Alumni and Contacts of those individuals that have gone through a W-2168/ASRA lab but now reside in Africa.

    Another source for international development in seed science might be the Consultative Group for International Agricultural Research (CGIAR).

    Commodity groups were also discussed and, in certain instances, can provide individual PIs with research funds. One example put forth was that Cotton Inc. has funded research in those states designated as a Cotton growing area. Fiber length and the high cost of cotton seed are two areas that are of interest to those growing the commodity. The Cotton Inc. web site highlights Agricultural Research that they are involved in through collaborative efforts with those in the public and private sectors (see http://www.cottoninc.com/biotechnology/).

    The American Seed Research Foundation would be another multi-state possibility to fund research in seed biology. It might be possible to spin to the companies paying in to the ASRF that part of the bad press they are getting for charging as much as they are for improved seed could be ameliorated if they advertise that they redirect a portion of this revenue back into seed research. If it would be possible to have a check-off for seeds sold, this would be a real and relevant source of funding to support Ph.D. student research to provide the seed industries with educated employees in the future.

    To this end, the NSF-GOALI (NSF-Grant Opportunities for Academic Liaison with Industry) program might just be an avenue to obtain funding to spearhead an effort to define an estimate of the personnel needs for the Seed industry in the USA for the coming decade. This would be a collaboration between Institutes and Companies. Then, with the numbers in hand, and the awareness of their predicament that their participation in the survey would bring, perhaps the ASTA would be more willing to forward the W-2168/ASRA objectives, seeing them clearly for their own.

    It was mentioned that Agricultural Economists are interested in assessing the needs of the Seed Industry perhaps we could get representatives from this group on board should a GOALI be deemed desirable.

    Regarding the outreach of the group, someone brought up the WIKI environment as an example of social networks acting as a learning environment. If such an effort was made by any of the sub-groups of the membership, the existing ASRA web-page would be an ideal site on which to host the pages.

    It was now after 3:00 pm and so it was moved that the meeting would be adjourned (Bruce) and seconded by Mitch McGrath. A vote was taken and passed unanimously.

    See minutes attachment.

    Impact Statements:
    1. The Ohio State University: Refinement of rapid systems to assess vegetable and other crop seed quality and vigor.
    2. Oregon State University: We provided knowledge on the involvement of microRNAs and their targets in seed germination and seedling establishment. Comprehensive understanding of molecular repression and de-repression by miRNA will aid in the development of seed germination technologies for the future. Characterization of AUXIN RESPONSE FACTOR10, a miRNA160 target, provided new insight into the crosstalk between auxin and ABA signal transduction, which is probably important for stress responses of seeds.
    3. Oregon State University: Undergraduate students gained skills in molecular, biochemical and genetic experiments. These students also authored grant proposals and gave presentations through their Howard Hughes medical Institute Summer Undergraduate Research grants. Four international seed scientists (Canada, India, Poland and Spain) joined the project and learned skills in seed research. A seed technologist from a seed company was trained and advanced his understanding of fundamental science.
    4. University of Florida: Conservation/restoration practitioners and seedling producers can better plan and coordinate in terms of seeding activities for restoration of degraded lands.
    5. Louisiana State University: The Spartina system permits separation of drying effects from those associated with recalcitrant seed death, providing a means to analyze seed parameters in the dormant state, independent of active seed metabolism. Oxidative stress is not the cause of recalcitrant seed death in Spartina suggesting that prior work on recalcitrant seed death has been compromised by technical and physiological artifacts. The long-term outcome of Spartina seed research will be the identification of the causes of recalcitrant seed death. This will provide the basis for generating desiccation-tolerant Spartina alterniflora seeds for commerce and genetic conservations efforts. Principles identified will enhance genetic conservation of all recalcitrant species.
    6. USDA-ARS-Michigan State University: An in-depth understanding of response to stress is critical during sugar beet seed germination in order to identify key germplasm and breeding lines for enhancing seedling vigor, and identifying mechanisms of seedling vigor under different germination regimes and in different genetic backgrounds. New germplasm was identified with tolerance to salt stress during germination. Manipulation of hormone sensitivity via selection and breeding allows rationale investigation of hormones in improving emergence and stand establishment in the field.
    7. University of Calfornia, Davis: This work has also involved collaborators or clientele in Regional Research Project W-2168, the NSF-funded Compositae Genome Project, the USDA-AFRI-NRI program, the UC Davis Genome Center, and the Western Regional Seed Physiology Research Group (a seed industry research support group), the American Seed Research Foundation, the American Seed Trade Association, the California Seed Association and the UC Davis Seed Biotechnology Center.
    8. Virginia Polytechnical University, Blacksburg: Demonstrated the potential for improving soil through preplant treatments with dilute polysaccharide solutions that stimulated soil microbial activity. Showed the importance of glutathione as a stress protectant in germinating and developing seeds. Trained 15 U.S. and international students in principles of vegetable seed production via distance learning over the world wide web.
    9. University of Kentucky: A full-day workshop, A short-course on seed dormancy and germination with emphasis on Hawaiian species University of Hawaii, Manoa, 31 July 2009. Half-day workshop, Future research objectives for studies on seed dormancy and germination in the Junggar Desert, northwest China. Xinjiang Agriculture University, Urumqi, China, 27 June 2009.
    10. The membership was expanded, an official name for the W-2168 that will not change each time we go through a re-write, was adopted, a webpage for the group was established.
    11. We have published our results adding to an emerging picture of direct transcriptional control. We have found, as have others, that while many sites are occupied by a given transcription factor in vivo (generally 1000s of DNA binding sites), relatively few interactions between a transcriptional regulator and DNA result in significant changes in gene expression of the nearby gene.
    12. Analysis of genes directly and indirectly controlled by AGL15 has led us to investigate hormone interactions involved in control of somatic embryo development. We are currently using combinations of mutants, hormones and inhibitor treatments to tease apart how different hormoneal cross-talk controls somatic embryo development. We have extended our work in Arabidopsis and demonstrated that ectopic expression of AGL15 in soybean can promote somatic embryogenesis in this important crop.
    13. University of Tennessee: Candidate genes showing differential expression in dry and germinating switchgrass seeds were identified. Some of these genes may be critical for regulating switchgrass seed dormancy and germination. Further functional characterization of these genes may lead to novel insights into the molecular mechanisms of seed dormancy and alleviation in switchgrass. Such information may provide an important knowledge base and tools for genetic improvement of switchgrass as a bioenergy crop.
    14. University of California Berkeley - Our work broke new ground in the following areas: (1) Cysteine biosynthesis in chloroplasts. Our results showed that a cyclophilin, CYP20-3, links photosynthetic electron transport and redox regulation to the folding of an enzyme of cysteine biosynthesis, SAT1, thereby enabling the cysteine-based thiol biosynthesis pathway to adjust to light and stress conditions. This was a longstanding unsolved problem.
    15. UC-Berkeley cont: (2) Chloroplast protein import. Experiments carried out jointly with the University of Munich provided the first evidence for a link of protein import to redox regulation via thioredoxin. Redox has long been known to regulate fundamental chloroplast processes. Our work extended this role to chloroplast protein import.
    16. UC-Berkeley cont: (3) Regulation of starch biosynthesis in chloroplasts and amyloplasts. We showed that the unusual plastid-localized NADP-thioredoxin reductase C (NTRC) containing both an NADP-thioredoxin reductase (NTR) and a thioredoxin regulates ADP-glucose pyrophoshorylase, the main regulatory enzyme in starch biosynthesis. The results provided biochemical and genetic evidence for a role of NTRC in regulating starch synthesis in response to either light or sucrose.
    17. UC-Berkeley cont: (4) The data also provided evidence that the thioredoxin domain of NTRC and, to a lesser extent, free thioredoxin linked to ferredoxin enable amyloplasts of distant sink tissues to sense light used in photosynthesis by leaf chloroplasts and adjust heterotrophic starch synthesis accordingly. The experiments have hopefully solved the mystery of how leaves communicate the light signal to roots.
    Last Modified: 09-Oct-2010

    Date of Annual Report: 08/09/2011

    Report Information:
  • Annual Meeting Dates: 07/08/11 to 07/09/11
  • Period the Report Covers: 07/2010 to 07/2011

  • Participants:
    Brief Summary of Minutes of Annual Meeting:
    8:35 a.m.

    Mark Bennett: Call to order and opening remarks.

    Introduction of participants.

    Mark: 1) Report on the state of W-2168 membership; 2) approval of 2010 minutes: approved by Daniel and Greg; 3) Outgoing leadership: Hector Perez (FL) is stepping up to President position; and Xingyou Gu (SD) is moving up to vice-president position; and 4) the next host site selection, plus nomination (secretary-elect) committee: Bob and Mark were appointed; Hector (FL) has offered to host the next annual meeting; time to be determined.

    Other topics discussed: Recruitment of new members - young faculty, graduate students, and research associates. Funding for graduate student travel. Budget considerations at individual institutions: CRIS or Hatch funding.

    Manjit: Introduction of Dr. Joe Colletti, Sr. Associate Dean of ISU College of Ag and Life Sciences.

    Dr. Colletti Statement of support: Ag Experiment station sees value in research collaboration. "Scope out the frontier--where should the science go?" Ethanol is 1st generation in the biofuel-economy; Soy is the 2nd, with oil used in biodiesel; The paradigm shift in agricultural industries; Dry distiller's grains no longer waste product, but a co-product, etc.; Working with the producers and public sector: Range of involvement = economists, engineers, scientists, producers, etc.; Funding for facilities is available.

    Questions for Dr. Colletti: "What examples have you seen to keep the seed science faculty and student positions [participation by industry]?" Joe: Recognize that if land grant institution programs thrive, Pioneer Hi-Bred et al. will benefit. Some opportunities present in an inorganic manner. Manjit: strong support, response to workshops, good interactions with industry. Joe: pay attention to the orphan crops. i.e., many apple orchards dropped out of production because of weather challenges, upswing in production of crops due to introductions of hybrids, etc. Resurgence of formerly-major crops now. Push for shipping seeds, not plants. Inclusion of smaller companies is important, too.

    Bret Hess (W-2168 Admin. Advisor):

    1) Name change for review committee (was RCIC, now MRC - multi-state rev. cmte.)

    2) Single submission deadline for ALL proposals, 15 January: New & renewals, Summer review, Approval during summer meetings, Fiscal year for federal govt. in October.

    3) Implication for W-2168: renewal must be submitted in 4th year; Engagement of stakeholders, industry reps, and inclusion of new members is acceptable; Important: must demonstrate progress for this to be acceptable for renewal of proposal.

    4) Advice: 10 July 2011 is date of next RMC. W-2168 is topic on agenda; our project seems to be stable; Review progress.

    5) Questions?

    a. "What about the next rewrite. The previous was quite extensive". Explanation of review process: Peer review, MRC review. Some projects overly ambitious. Streamline proposals to minimize rewrites. Need to submit a report on W-2168 progress.

    b. "Since you have been involved..submissions that are not accepted..is the bar raised?" Can't recall any declined outright. Most are revised first. Governance outline is not necessary&be mindful of what is required. This group has been successful, so no problems anticipated. Some advice may be provided to this group. Proper use of terminology, 'milestones', 'outcomes' vs 'outputs', etc.

    c. "Is there a national project?" This is considered a national project. This group is based in western region, but the projects are open throughout the region. It just happened to land here. Not sure how that happened- likely linked to extensive seed production/research activity on west coast years ago (and ongoing). The higher level function is not filled by this group, which is a research group. The reason for the western region is that this group started in the western states originally. As other seed people from other states joined or left the group, the shift in group locations shifted over time. Some of the things that we seem to have excelled in are educational programs. We can discuss how industry sees us as a pipeline.

    d. Research is the first priority for this group, but should we tout the number of students as output for this group? RMC & include outreach as part of an objective. RMC is starting to see a trend in a formally-structured education objective, either through outreach/extension or education. Some groups would invite stakeholders to participate in group, i.e., invite Pioneer to W-2168 meetings. The more broad, the more global, the better the reception [to this proposal].

    e. "The new project..will we need to prepare the proposal for the next project in 2012 or 2013?" Prepare in 2012, so if revisions are required, there is time to revise. Otherwise, the 2013 deadline may be missed, funding not granted, and this group runs the risk of being inactive.

    f. Final advice: the report for this meeting is due within 60 days (September 8). Remember the form indications are limited, so streamline the report. Last year's report was too lengthy.

    g. "How do we update our participation list?" Give Bret a list, so he may contact their AES for inclusion or removal, or work with your institutional AES to update that information.

    h. Even if each institution supports only 1 designated official member, it would be acceptable to bring additional representatives but keep the 1 designated member. Representation of each state from more than one institution would be favorably viewed (i.e., one rep from each land-grant, one rep from an additional univ. or agency).

    Introduction of new attendees.

    Daniel: Brief introduction of South American and African countries contacts.

    10-minute break, then state reports.

    State report:

    1. Bob Geneve (KY): 1) Overview of UK group: Bruce Downie , preparing for sabbatical leave; Jerry Baskin has retired; Carol will retire soon. 2) Research report on searching for the water gap in seeds with physical dormancy.

    2. Xingyou Gu (SD): 1) Genetic and QTL analysis of seed dormancy and pre-harvest sprouting in synthetic wheat; and 2) cloning and molecular characterization of QTLs for seed dormancy rice.

    3. Lin, Liang-Shiou (USDA-NIFA): Introduction of NIFA structure; 5 challenging areas. Questions: NIFA collaboration with NIH and other funding agencies.

    4. Greg Welbaum (Virginia Tech): BARD project: control of bacterial fruit blotch of cucurbits using the maize non-host disease resistance gene Rxo1. BARD funded by Israel. BFB Bacterial fruit blotch caused by Acidovorax avenae. 600 lines of watermelon identified for responses to the disease. Cold tolerance: base germination temperature of Cantaloupe seeds. Drying seeds using clay beads for tropical crops.


    Tour laboratories of Seed Sci. Center.

    Afternoon state report:

    5. Lisa Shepherd (ISU): Intro. of National Seed Health System (NSHS) & phytosanitary efforts.

    6. Gary Munkvold (ISU): NSHS, methods development. Seed borne diseases in corn and soybean. Graduate program in seed tech and business. Online delivery of teaching programs.

    7. Daniel I. Leskovar (Texas A&M): Improving transplant quality and stand establishment of vegetable transplants under biostress. Does ABA protect watermelon seedling from chilling injury when transplanted in early spring? Stress tolerance of pepper (Colima)-leaf chlorophyll. 2) ABA: height control on pepper and watermelon. ABA staging expt. with watermelon. Crop protection from deer damage. Yellow nutsedge: chufa.

    8. Hiro Nonogaki (Oreg.St.): a) Hormone metabolism in seeds; plant gene switch system (PGSS), promoter+NCED6 induced by chemical. Ligand:methoxyfenozide NCED6 induction; ABA deactivation system or GA biosynthesis GA3 ox1. ABA perceptation and signaling. Seeds III - Coming soon.

    July 9, 2011

    8:27 am: Call to order. Distribution of contact information. Final points before beginning today's business.

    State reports (cont.)

    9. Peggy Lemaux (CA):

    1) Brief report on accomplishments from Kent Bradford's group: Identification and characterization of biophysical, biochemical, genetic, and environmental factors regulating or influencing seed development, germination, vigor, and dormancy in lettuce. LsNCED4 is essential for induction of thermo-inhibition in lettuce seeds.

    2) Modifying expression of thioredoxin in cereal seeds to improve grain properties including PHS (UC-Berkley).

    10. Alan Taylor (Cornell Univ.- Geneva): Seed dormancy in switchgrass; Seed sci & tech controlled release of seed treatments; seed conditioning of warm season grasses; introduction of rotary pan coating tech. Microcrystalline cellulose infused with Rhodamine B as colored and fluorescent tracer.

    11. Keting Chen (Graduate student in ISU Seed Sci. Center): Cellular mechanisms of seed osmopriming in spinach.

    12. Mark Bennett (Ohio St.): Relationships between internal morphology (esp. % free space), germination and vigor of Portulaca grandiflora and Portulaca sp seeds, using non-destructive x-rays and scanning software

    Final discussion:

    1)Site selection for the next W-2168 annual meeting. New president (and host?): Hector Perez (FL); time: Jan, Feb or Mar. 2012 ; survey membership for best dates; location: pending details from Hector.

    2)Report of secretary-elect - Manjit will recruit for position from ISU colleagues.

    3)Free discussion:

    Manjit: Distribution of presentation list/speaker schedule before the conference; make the meeting available to graduate students; get industries & ARS people involved in the meeting.

    Greg: W-2168 symposium every ~ 4 years to have industries involved.

    Manjit: invite other seed groups to the meeting.

    Bob: Feedback from seed industries for new issues to be addressed by seed researchers.

    Manjit: milestone accomplishments, add/combine symposium to W-2168 annual meeting.

    ISU seed sci. 2012 spring (and annually) symposium.

    Alan: Recruit young seed researchers to W-2168.

    Bob: Invasive species researchers; Germplasm groups.

    Greg: Proposal of 'seed CAP', potential areas such as seed health. Match funding for seed pathology from other industry partners?

    Lin: This community is a stake holder (inclusion of seed develop/health/biology in USDA NIFA RFA)

    Manjit/Peggy: additional seed sci. education opportunities (e.g., games/web info) to high school students.

    Adjourn at 11:00am

    Objective 1. Identify and characterize biophysical, biochemical, genetic, and environmental factors regulation or influencing seed development, germination, vigor and dormancy.

    Dr. Pablo Jourdan continues as Director of the revitalized Ornamental Plant Germplasm Center (OPGC), a partnership between the USDA and Ohio State University. The OPGC's research program explores factors that influence long-term seed storability for germplasm preservation. Peter Zale (PhD student) is using the core collection of 243 Phlox accessions to study seed biology potential for interspecific hybridization. (OH)

    We have characterized a major quantitative trait locus (QTL) termed Htg6.1 associated with the ability for lettuce (LACTUCA SATIVA) seeds to germinate at high temperatures (up to 37 C). Genetic mapping identified a candidate gene (LsNCED4), which encodes an enzyme in the abscisic acid (ABA) biosynthetic pathway (9-cis-epoxycarotinoid dioxygenase 4). Functional complementation of Arabidopsis mutants in homologous genes demonstrated that LsNCED4 has the expected biosynthetic activity. Lines having mutations in LsNCED4 were isolated and their seeds were able to germinate at much higher temperatures than their isogenic control lines. Silencing of LsNCED4 in a thermosensitive line of lettuce (cv Salinas) using RNAi enabled seeds to germinate at temperatures up to 40ºC. Transferring the promoter and coding region of LsNCED4 from a thermosensitive line into a thermotolerant line resulted in the induction of thermoinhibition in seeds of the latter. These results confirm that LsNCED4 is essential for the induction of thermoinhibition in lettuce seeds and that reducing the expression of this gene can enable germination at high temperatures. Transcriptomic analyses identified clusters or modules of genes that exhibit correlated expression patterns in association with high temperature inhibition of germination. A second RIL population is being mapped to identify additional Htg QTL. A strong QTL on chromosome 9 (Htg9.1) is being fine-mapped to pinpoint candidate genes. (CA - Davis)

    This project has supported research to identify the causes of seed dormancy and seed desiccation intolerance (recalcitrance) of the marsh grass, Spartina alterniflora, which is an important component for successful maintenance and remediation of Louisiana coastal stabilization. Two manuscripts were prepared and submitted; one has been accepted for publication and the other is in the process of peer review. New data have been collated in preparation for two forthcoming scientific meeting abstracts/presentations. The results obtained have been incorporated into lecture materials for undergraduate and graduate level courses, presented each year. The project has also supported the research training of one doctoral student (seed physiology, biochemical techniques, and mass spectroscopy) and one undergraduate (seed physiology and scientific method). Services included advising faculty and students in the LSU AgCenter Coastal Plants Project concerning development of seed germination protocols for various marsh plant species. (LA)

    Bimolecular fluorescent complementation and protein pull-down assays identified in vivo interaction between PHYTOCHROME INTERACTING FACTOR1 (PIF1) and COLD TEMPERATURE GERMINATING10 (CTG10). (KY)

    Using phage display and biopanning to identify protein interacting partners of PHYTOCHROME INTERACTING FACTOR1 (PIF1) bound to the promoter of GA INSENSITIVE (GAI). (KY)

    Identification of the water gap mechanism in Geranium carolinianum. (KY)

    Continued investigation of direct targets of the MADS-domain protein AGL15 in control of somatic embryogenesis has identified components of ethylene and auxin biosynthesis and response on promotion of somatic embryogenesis. (KY)

    Development of tools and techniques to identify direct targets of FUSCA3 and ABI3, two transcription factors important for seed development. (KY)

    Generation of transgenic Glycine max ectopically expressing MADS-box genes to test effect on promotion of somatic embryogenesis in this species. (KY)

    Identify quantitative trait loci (QTL) associated with seed dormancy and pre-harvesting in a doubled haploid (DH) population derived from synthetic hexaploid wheat. We developed a linkage map consisting of 285 simple sequence repeat markers on the 21 chromosomes of common wheat based on a population of 186 DH lines. With this framework map, we identified 19 QTL associated with seed dormancy and/or sprouting rate of intact seeds on spikes, which were evaluated for the mapping population grown under the local environment conditions from 2006 to 2010. Several QTL expressed in multiple years during the research period and could be used to improve local cultivars for resistance to pre-harvest sprouting. (SD)

    Clone and characterize qSD12, a major QTL for seed dormancy in rice. Five candidate genes in the narrowed qSD12 region were sequenced from donor and recipient parents of the dormancy QTL enhancing alleles and sequences annotated. More than 20 new markers were developed based on polymorphism between genomic DNA sequences from the two parents and the markers are been using to conduct a haplotype analysis for about 200 cultivars to understand the origin, differentiation and distribution of the major dormancy gene. Three of the five candidates have been transformed into a non-dormant cultivar to confirm their dormancy functions by complementation. Development of transgenic rice and transgenic plant analysis are under way. (SD)

    Identify downstream gene networks regulated by SD7-1, a seed dormancy gene cloned from weedy rice. SD7-1 was map-based cloned as the red pericarp color gene Rc in weedy red rice and is annotated as a bHLH family transcription factor. Both microarray and yeast-two-hybridization (Y2H) analyses were used to identify gene networks regulating the seed dormancy and pigment traits and partners of the transcription factor. Three genes for biosynthesis of abscisic acid (ABA, a dormancy-inducing hormone) and nine genes in a conserved flavonoid biosynthetic pathway were up-regulated by SD7-1. Higher ABA content was detected in the dormant than in the non-dormant isogenic lines of SD7-1. Histological analysis demonstrated that the flavonoid pigment expresses only in the lower epidermal cell layer of the pericarp tissue. Y2H analysis is under way.(SD)

    Develop isogenic lines for additional seed dormancy QTL. Seed dormancy QTL qSD1-1, 1-2, 4, and 7-2 loci were finely mapped onto the genomic region of <100 kilo base pairs and the dormancy-enhancing alleles introduced into the same genetic background to facilitate cloning and molecular characterization of the QTL underlying genes. (SD)

    Since our last report, we have focused on improving wheat quality through changes in the expression of genes for thioredoxin (Trx) - regulatory protein functional through biology. The results suggest that by changing the levels of this key protein, wheat can be improved in two fundamental parameters: allergenicity and preharvest sprouting. The work has been published in comprehensive papers, coauthored with U.S. and Chinese collaborators. (CA, Berkeley)

    Work with cereals (barley and wheat) and a legume (Medicago truncatula) has established extraplastidic Trx h as a central regulatory protein of seeds. Trx h acts by reducing disulfide (S-S) groups of diverse seed proteins (storage proteins, enzymes and enzyme inhibitors) thereby facilitating germination. Early in vitro protein studies using addition of components of the Trx system were complemented with experiments in which barley seeds with Trx h overexpressed in the endosperm showed accelerated germination and early or enhanced expression of associated enzymes (alpha-amylase and pullulanase). The current study extends the transgenic work to wheat. In the present study, [1], two approaches were followed to alter the expression of the Trx h genes in the endosperm: (1) a hordein promoter and its protein body targeting sequence led to overexpression of Trx h5, and (2) an antisense construct of Trx h9 resulted in cytosolic underexpression of that gene (Arabidopsis designation). Underexpression of Trx h9 led to effects opposite to those observed for overexpression Trx h5 in barley - retardation of germination and delayed or reduced expression of associated enzymes. Similar enzyme changes were observed in developing seeds. The lines with under expressed Trx showed delayed preharvest sprouting when grown in the greenhouse or field without a decrease in final yield. Wheat with overexpressed Trx h5 showed changes commensurate with earlier in vitro work: increased solubility of disulfide proteins and lower allergenicity of the gliadin fraction. The results are further evidence that the level of Trx h in cereal endosperm determines fundamental properties as well as potential applications of the seed. (CA - Berkeley)

    In a more recent study [2], we obtained evidence that Trx h9, which was found to be bound to the plasma membrane, acts in a manner not previously seen for a Trx-i.e., it can move from cell-to-cell. Analysis of Trx h9 revealed a 17-amino acid N-terminal extension in which the second Gly (Gly2) and fourth cysteine (Cys4) were highly conserved. Mutagenesis experiments demonstrated that Gly2 was required for membrane binding, possibly via myristolation. Both Gly2 and Cys4 were needed for movement, the latter seemingly for protein structure and palmitoylation. A three-dimensional model was consistent with these predictions as well as with earlier evidence showing that a poplar ortholog of Trx h9 is reduced by a glutaredoxin rather than NADP-thioredoxin reductase, the reducing partner of other Trx's. In demonstrating the membrane location and intercellular mobility of Trx h9, the present results extend the known boundaries of Trx and suggest a role of Trx h9 in cell-to-cell communication. This work, which we plan to follow up with wheat, gives a new perspective to redox regulation and its role in seed development and germination. (CA - Berkeley)

    Sugar beet is tolerant of high salinity, except during germination and stand establishment. Selection for improved germination in saline solutions was accomplished. Seed from one of these selections, EL-A022799 derived from Ames 3051, was evaluated using LC/MS/MS during germination in order to examine the presence and prevalence of small molecules implicated in salt tolerance in other species. Seedlings germinated in 0, 75, or 150 mM NaCl were evaluated for levels of glycine betaine, proline betaine, betaine aldehyde, choline, proline, sarcosine, and azetidine-2-carboxylic acid. Proline betaine was found at low levels in seedlings germinated in water, but the concentration of this compound decreased to barely detectable in salt germinated seedlings. Glycine betaine was abundant in the salt-selected Ames 3051 germinated in water, and was barely detectable in the non-salt tolerant control in the same conditions (and which did not germinate in NaCl). Interestingly, the glycine betaine level in salt-selected Ames 3051 decreased in response to increasing salt content in the germination medium until it reached a level similar to the control at zero salt, after which germination was not effective. (MI)

    A candidate gene transcript profiling was untaken to examine early growth stages during germination. Genes involved in stress responses, signaling, and growth were activated within one hour after imbibition. Gene expression patterns were remarkable dissimilar between two contrasting legacy hybrids over the first 24 hours of germination. Fruit and true seed water uptake was measured in water and in hydrogen peroxide, with the result that hydrogen peroxide, which stimulates germination, does not appreciably affect fruit characteristics but rather acts on the true seed. Candidate genes demonstrated the involvement of a number of biochemical processes in the expression of seedling vigor, and some of these are still evident in contributing vigor in three-week old seedlings. (MI)

    Tomato GeneChip analysis was performed to identify tissue-specific genes in tomato seeds. Tomato seeds were imbibed for 18 h and then dissected into the micropylar part and the rest of seed (termed as 'lateral' part). The embryonic tissues were removed from the micropylar and lateral parts, which were named endosperm cap (EC) and lateral endosperm (LE), respectively. Although both parts still contained testa (seed coat), the term endosperm was used because the testa is non-viable tissue in the mature tomato seed and does not affect expression analysis, an objective of which was to compare endosperm- and embryo-specific gene expression. The embryo was also divided into radicle- and cotyledon halves, which were named radicle-half embryo (R) and cotyledon-half embryo (C), respectively. RNA extracted from these four tissues was used for GeneChip analysis. EC-specific genes were identified, which included cell wall-associated genes, pathogenesis-related (PR) genes, and hormone metabolisms genes. (OR)

    Analysis of the promoter regions of the EC-specific PR genes indicated that ethylene response factors might be the upstream regulators of the EC-specific genes. Consistently, tomato ETHYLENE RESPONSE FACTOR 1 (TERF1) was detected as one of the EC-specific genes. Bioinformatic analysis indicated that the promoter regions of the cell wall-associated genes, such as mannanase and expansins, also contained DNA motifs that are recognized by ethylene response factors. These results opened the possibility that ethylene response factors are the major upstream regulators of many genes specifically induced in the endosperm cap of tomato seeds. (OR)

    Hormone Metabolism in Seeds. Genes encoding hormone metabolism enzymes that are important for seed dormancy and germination, such as abscisic acid (ABA) and gibberellin (GA) biosynthesis and deactivation enzymes, have been isolated. These genes are though to be controlled mainly at the level of transcription. A single transcriptional change in any of these genes could alter hormone levels in seeds and seed performance in terms of germination and dormancy. To enable manipulation of these rate-limiting enzymes, Plant Gene Switch System, an inducible gene expression system has been applied. Induction of an ABA biosynthesis gene was able to increase ABA levels in seed significantly and caused near-complete suppression of germination, providing a proof of concept for hormone metabolism engineering for seeds. (OR)

    Relationships were developed with seed producers in the emerging wildflower seed industry of the southeast US and restoration/conservation practitioners to identify key areas of seed biology research. This is evidenced by $180,336 in funded grants spanning 200912. Funded projects revolved around seed dormancy in pre-variety germplasm and its alleviation, germination phenology, genetic diversity among seed producing populations of keystone species, development of viability testing methods, soil seed bank formation, and cryopreservation of wild germplasm. (FL)

    Muskmelon (Cucumis melo L.) is a warm-season, frost-sensitive crop that germinates poorly at low temperatures. Cold-sensitive cultivar Noy Yizre'el (NY) and cold-tolerant cultivar Persia 202 (P-202) were compared at 15 and 25 degree C to determine how seed coat anatomy and oxygen uptake affected low-temperature germinability. The seed coats of both cultivars restricted oxygen uptake but 'NY' had a lower rate of oxygen uptake compared to P-202 with or without the seed coat. The poor germination of NY at lower temperatures was related primarily to lower embryo oxygen uptake in addition to the seed coat restriction of oxygen uptake. (VA)

    Switchgrass (Panicum virgatum L.) is used horticulturally as an ornamental and agronomically as an animal feedstock and a putative bio-energy crop. Genetic transformation, using somatic embryogenic (SE) callus derived from mature seeds, is one strategy for improving switchgrass traits. A superior switchgrass line, HR8, was developed in using recurrent tissue culture selection from cv. Alamo that has low dormancy and produces a high percentage of somatic embryos. Eighty two percent of HR8 seeds germinated after harvest comparing to 26.8% for unselected Alamo. HR8 seeds that germinated produced 84.9% SE callus. (VA)

    Evaluated Lycium chinense, Chinese goji berry, production from plants developed from tissue culture protocol developed in collaboration with Institute for Advanced Learning and Research Danville, VA. (VA)

    Basil is a popular herb but little is known about its base, ceiling and optimal germination temperatures. Characterized the germination performance of two basil seed lots at different temperatures. (VA) Thermogradient tables are used to generate a wide range of temperatures for seed testing on the surface of an aluminum plate. A new thermogradient design with a finned surface to maintain temperatures vertically was developed. (VA)

    Objective 2. Determine and model the biotic and abiotic factors affecting seed germination, seedling emergence, and establishment of sustainable populations in natural and agro-ecological systems.

    We are collaborating in a workgroup on 'Germination, Trait Coevolution, and Niche Limits in Changing Environments' sponsored by the National Evolutionary Synthesis Center (NESCent) at Duke University. Though this group, we are working with colleagues at the University of Arizona (Larry Venable) and at the Institute of Botany, Chinese Academy of Sciences (Zhenying Huang) to conduct hydrothermal time analyses of seed germination of 13 native or introduced species. Dr. Venable has over 25 years of data on germination of these species in response to seasonal changes in temperature and rainfall, and we are analyzing their hydrothermal time responses in relation to after-ripening to assess how changes in dormancy relate to their ecological fitness and demographic success. We are also providing input to other members of this group on incorporating hydrothermal time concepts into demographic population models attempting to model and predict behavior of seed populations under changing environmental conditions, dependent upon their phonological life histories. (CA - Davis)

    Used phage display and biopanning to identify PROTEIN ISOASPARTYL METHYLTRANSFERASE (PIMT) targets in Arabidopsis. (KY)

    Using phage display and biopanning to examine the Arabidopsis LEA protein, SEED MATURATION PROTEIN1 (SMP1), and the soybean homolog, GmPM28, for protein binding partners at high temperature. (KY)

    Objective 3. Develop, evaluate, and transfer technologies to assess and improve seed and seedling quality, health, performance, utilization, and preservation.

    Research at Ohio State Univ. by Vanessa Neumann Silva, USP-ESALQ (Brazil) studied the relationship between Portulaca grandiflora and Portulaca sp. (hybrid) seed morphology, germination and vigor. Five lots of Portulaca grandiflora and two lots of Portulaca sp. were analyzed by non-destructive x-ray testing and subsequent germination, accelerated aging and seedling computer analysis by the Seed Vigor Imaging System (SVIS®). We verified the frequency of occurrence of free space for each lot. Accelerated aging data and image analyses indexes were submitted to ANOVA and Tukey's test with 5% of probability. The P. grandiflora seeds had free space between the embryo and endosperm ranging from 0-4.5%, in relationship to the whole area of the seed, and more than 8-0% of these seeds had free space between 0-1.5% for all lots. For lots 2, 3 and 4 the increasing free space was directly proportional to reductions in germination. From accelerated aging test data and seedling image analysis, lots 2 and 3 were the most vigorous; however, in these lots the percentage of seeds with extended free space (1.5-4.5%) was similar to the other lots, which suggests that in this situation free space had no influence on seed vigor. For Portulaca sp. lot 6 had large amounts of free space (almost 10% in class 3), and had lower germination, lower SVIS vigor index values and seedling length, lower germination after accelerated aging, and lower indexes of vigor, growth and uniformity of growth after the 72h of accelerated aging in relationship to lot 7. (OH)

    The respiratory activity of individual seeds during imbibition and germination is being assessed using the Q2 instrument from ASTEC Global. Initial seed respiration during imbibition and germination is being characterized in relation to seed vigor and the effects of temperature and water potential on germination. The effects of respiratory inhibitors are also being tested to identify the metabolic pathways underlying different components of the oxygen uptake patterns exhibited by imbibed and germinating seeds. (CA - Davis)

    The effects of seed drying methods on seed longevity are being assessed. Seeds exhibit hysteresis in their moisture content/relative humidity relationships, or isotherms. Seeds that are drying have moisture contents associated with a desorption isotherm this is generally 0.5 to 1% higher in moisture content than seeds that are hydrating to the same equilibrium RH on their sorption isotherm. This difference in seed moisture content results in differences in seed longevity. Based on these results, simple modifications to seed drying procedures may be able to extend seed longevity in storage. (CA - Davis)

    With funding from the USAID-funded Horticulture Cooperative Research Support Program (HortCRSP), we are demonstrating and disseminating desiccant drying bead technology for seed drying and storage by smallholder farmers in India and Nepal. Seed drying beads can dry seeds efficiently in hermetic containers, and can be regenerated by heat and reused indefinitely. This technology has considerable promise for improving seed quality in humid regions. (CA - Davis)

    It was determined that for gamagrass applications that require an immediate, specific field population, purchasing decisions and planting rates should be based on Pure Actual Germination (PAG), calculated from seed lot purity and actual germination percentages rather than a Pure Live Seed (PLS) basis. (KY)

    Abscisic acid (ABA) treatments (frequency, rates, and timing of application) were investigated to reduce drought stress in jalapeno cv. Colima and bell pepper cv. Excursion and in watermelon cv. SF 800 (seeded) and SS 5244 (seedless). ABA was also studied as a potential tool to reduce chilling stress in seeded cv. Stargazer and seedless cv. Majestic watermelons in both greenhouse and immediately after field transplanting. (TX)

    We conducted a project to address the accuracy of seed health tests on P. stewartii in corn and found that all of the available PCR methods will cross-react with other species of Pantoea. In addition, several unidentified isolates of Pantoea spp. have been isolated from corn in several countries, and we are investigating the genetic relatedness of the unknown species with known species of Pantoea. Currently we cannot recommend the use of PCR-based tests for P. stewartii phytosanitary testing. (IA)

    Soybean seedborne pathogens - We completed a project in 2011 investigating interactions between seedborne viruses (SMV and BPMV) and Phomopsis longicolla. We found that infection of soybean plants by BPMV increased the susceptibility of these plants to seed infection by P. longicolla, and this predisposition was not related to virus effects on plant developmental rates. SMV did not have a similar effect. When vectors of BPMV are active, controlling them can contribute to control of both BPMV and seed infection by P. longicolla. (IA)

    Corn seedling pathogens and nematode interactions - We are studying the interactions between root-lesion nematodes and Fusarium verticillioides, Fusarium graminearum, Rhizoctonia solani, and Pythium ultimum. Seedling disease symptoms were enhanced when nematodes were present in combination with any of the fungi, but the strongest interactions occurred with F. verticillioides and R. solani. Seed treatment combinations, including several fungicides and a nematicide (abamectin), provided the best improvements in seedling root development and growth. (IA)

    Several new projects on soybean seed storage and drying are underway, as well as studies on soybean epigenetics and seed quality. Preliminary results on seed storage showed a poor relationship between excessive soil nutrients availability during plant growth and seed survival in storage. However, excess nutrients can be detrimental to seed quality. Seed priming can enhance epigenetic changes in the agronomic performance of the plant. (IA)

    Work on mapping QTLs for germination of maize under low temperatures continues. We are investigating whether QTLs are expressed differently based upon the temperature of the stress conditions, and we have another study planned for this fall. (IA)

    A flow meter was developed to measure the flow continuously and in real time for seed operations. A prototype was built and installed in a seed plant in Williams, Iowa. The data collected from Remington Seeds' Williams plant was analyzed and both electrical and mechanic vibration noise was observed. Modification of load cell mounting and electric isolation solved the problems. The prototype then was installed in the foundation seed plant of the CAD (Committee for Agricultural Development) in Ames for further testing. The flowmeter data at CAD was collected and analyzed. The plant manager reported that the flowmeter produces much less dust and is quieter. No left-over seeds were found inside the flowmeter and no abnormal wearing was observed on both the sensing unit and rubber boots. Test results indicated high corrections (r2=0.9909, 0.9903, and 0.9719 respectively for Large Flat corn, small round corn, and soybeans) between the measure weights and PLC readings. (IA)

    Taylor's lab has state-of-the art seed coating technology using rotary pan technology, and he is a cooperator with pest management specialists at and outside Cornell. New chemistry seed treatments were applied to vegetable crops for control of insects or diseases. (NY)

    Switchgrass (Panicum virgatum) is being explored as a biofuel crop, and weed management is a major production problem during the first year. Our approach was to apply an herbicide safener as a seed treatment to protect of the seed and seedlings from potential phytotoxicity from a chemical herbicide. (NY)

    Bacteria Fruit Blotch (BFB) is a seed transmitted disease of cucurbits caused by Acidovorax avenae subsp citrulli (Aac) that is one of the most devastating diseases that affects both watermelon (Citrullus lanatus) and cantaloupe (Cucumis melo) production worldwide. Characterization of Aac strains revealed the existence of at least two well-differentiated groups of the disease. The first group includes strains that were mainly isolated from cantaloupe and pumpkin. While the second group represents the typical BFB strains that affect watermelon. The genome of Aac strain AAC00-1 (group II) was sequenced. Annotation of Aac genome revealed the existence of eleven putative type III effector genes including an avrRxo1 homolog. We cloned all known type III effectors from both group I and group II Aac strains and transiently expressed the effectors in a collection of cucurbit germplasm and Nicotiana benthamiana plants by using a Agrobacterium-mediated transient assays. The screening led to the identification of cucurbit germplasm that responds to the expression of Aac type III effectors. We plan to screen more cucurbit germplasm for resistance to specific effects, characterize the interplay of different type III effectors, and test which effectors have significant virulent contribution of Aac. (VA)

    Recent human disease outbreaks of bacterial pathogens transmitted on vegetables, especially fresh spinach, have raised questions about the potential for seed transmission of human pathogens. The bacterial populations of spinach seed and seedling leaves were compared using DGGE, to assess bacterial community richness, and real-time PCR to compare the abundance of select phyla (total bacteria, Actinobacteria, Bacteroidetes, Firmicutes, a-Proteobacteria and b- Proteobacteria). To determine the effect of environment, the plants were grown in field and growth chambers. Vertical transmission of bacterial community members was evident because the developmental stage of the plant affected the richness and abundance of select bacterial phyla. The bacterial richness of plants grown in the two environments was not affected. However, a greater number of bacteria were associated with field grown samples in comparison to those produced in growth chambers. A statistically significant interaction existed between growth stage and environment for each of the selected phyla. Populations on cotyledons were smaller than mature leaves, but were not significantly different than the 3-4 leaf stage seedlings. The culturable populations of bacteria on seeds (~5 log CFU/g) were significantly smaller than determined using real time PCR (~7 log copies). Vertical transmission of bacteria from seeds to seedlings, suggests that bacteria may have evolved to ensure the transmission of certain bacteria. (VA)

    Impact Statements:
    1. Ohio State University: Refinement of rapid systems to assess vegetable, flower and other crop seed quality and vigor.
    2. University of California - Davis: Mutants in LsNCED4 that exhibit ability to germinate at high temperature can be made available for integration into lettuce cultivars. This should help alleviate thermoinhibition and improve efficiency of lettuce production in desert growing areas. Drying beads can be utilized in many scalable applications for drying seeds and other horticultural products.
    3. Louisiana State University: Over 100 differentially expressed proteins among Spartina alterniflora (desiccation intolerant, DI), Spartina pectinata (desiccation tolerant, DT) and Spartina spartinae (desiccation tolerant) seeds were found by two-dimensional gel electrophoresis between DT vs. DI species, and some of these differences included differential post-translational protein phosphorylation and protein glycosylation. Some of these protein profile differences were detected after fractionation of total soluble protein extracts, while others were detected in a heat-stable protein fraction. Spartina alterniflora seed protein profiles also changed as a result of storage of the seeds in cold water, which we have shown triggers the transition from the dormant to the germinable, non-dormant state. The identification of these proteins via mass spectroscopy should provide clues as to the nature of desiccation intolerance and dormancy of Spartina alterniflora seeds.
    4. South Dakota State University: The wheat DH lines with the information on marker genotypes and seed dormancy QTL could be selected to enter wheat breeding programs. The newly selected isogenic lines for seed dormancy alleles introduced from weedy rice can be used as breeding materials to manipulate germination capability of cultivars.
    5. University of California - Berkeley: Our wheat work has demonstrated that changes in the expression of different form of the regulatory protein, thioredoxin, can improve nutritional and agronomic properties of wheat. The work provides a foundation for further work by molecular biologist and plant breeders to bring these transgenic grains to the marketplace. The work also opens a new door to our understanding of redox regulation and its role in seed development and germination.
    6. Texas A&M University: In conjunction with the industry, we are developing and optimizing hormonal application methods to improve transplant quality while suppressing stem elongation rates in the nursery and to mitigate the negative effects of transplant shock that lead to poor stand establishment and crop performance in diverse field environments.
    7. Iowa State University: The science-based seed health tests are instrumental in protecting the export of American seed that is estimated at $800 M annually.
    8. University of Florida: The impact of our research is that conservation/restoration practitioners and seedling producers can better plan and coordinate in terms of seeding activities for restoration of degraded lands.
    9. Cornell University: Two materials received a federal label as onion seed treatments. Sepresto contains two neonicotinoid active ingredients, clothianidin and imidacloprid, and is a product of Bayer CropScience. Entrust contains the active ingredient spinosad and is a product of Dow AgroSciences and will be marketed by Syngenta Crop Protection. Entrust is listed among the products approved by the Organic Materials Review Institute (OMRI) and can be used for protecting organically grown vegetable crops.
    10. Virginia Tech: Demonstrated a correlation between seed respiration rates and base temperature germination that could help plant breeders decrease the minimum temperature for cantaloupe seed germination. A switchgrass line, HR8, was developed using recurrent tissue culture selection from cv. Alamo. HR8 has low seed dormancy and produces a high percentage of somatic embryos from mature seeds for genetic transformation of switchgrass using somatic embryogenic (SE) callus. Supplied thermogradient tables to aid the seed research of colleagues. Trained 12 US and international students in principles of vegetable seed production via distance learning over the world-wide-web.
    Last Modified: 10-Aug-2011

    Date of Annual Report: 05/01/2012

    Report Information:
  • Annual Meeting Dates: 03/09/12 to 03/10/12
  • Period the Report Covers: 10/2010 to 09/2011

  • Participants:
    Brief Summary of Minutes of Annual Meeting:
    Accomplishments: Objective 1. Identify and characterize biophysical, biochemical, genetic, and environmental factors regulating or influencing seed development, germination, vigor and dormancy.

    A graduate student, Steven Haba, has begun a project involved with seed production and characterization in Begonia. This effort includes development of rapid and efficient quantitative seed germination tests for the very tiny seeds; assessment of basic biochemical properties (carbohydrate, oil, protein content); desiccation tolerance (seeds of some species appear to be orthodox and relatively long-lived); and development of accelerated aging protocols for evaluation of vigor and prediction of long-term storage. Selected accessions of Rudbeckia fulgida are being grown during 2011 to produce fresh seed and begin a more detailed comparative study of GA treatment, and to combinations of GA and cold among the accessions. We are developing approaches for production of at least 10,000 seeds per accession of species that tend to flower over a long time and have a concurrent long fruit ripening period, that upon maturity results in ballistic dispersal of the 1 or 2 seeds per fruit.

    Muskmelon (Cucumis melo L.) is a warm-season, frost-sensitive crop that germinates poorly at low temperatures. Cold-sensitive cultivar Noy Yizre el (NY) and cold-tolerant cultivar Persia 202 (P-202) were compared at 15 and 25 C to determine how seed coat anatomy and oxygen uptake affected low-temperature germinability. The seed coats of both cultivars restricted oxygen uptake but NY had a lower rate of oxygen uptake compared to 'P-202' with or without the seed coat. The poor germination of NY at lower temperatures was related primarily to lower embryo oxygen uptake in addition to the seed coat restriction of oxygen uptake.

    The qSD1-2 and qSD7-2 seed dormancy QTLs were narrowed to short genomic regions of several candidate genes and the two QTLs were also associated plant height. The fine mapping data suggest that qSD12 may consists of more than one underlying gene. Some of the putative QTLs for seed dormancy or resistance to pre-harvest sprouting in wheat detected in the previous years were confirmed with the new data and the map positions were improved. Three QTL alleles enhancing resistance to pre-harvest sprouting were introduced to the same genetic background.

    We have demonstrated using functional gene transfer and RNAi silencing that a major quantitative trait locus (QTL) termed Htg6.1 associated with the ability for lettuce (Lactuca sativa) seeds to germinate at high temperatures (up to 37 C) is due to an allele of a gene (LsNCED4), which encodes an enzyme in the abscisic acid (ABA) biosynthetic pathway (9-cis-epoxycarotinoid dioxygenase 4). Lines having mutations in LsNCED4 were characterized and exhibit thermo-tolerance during germination. These lines are being backcrossed to purify from additional mutations and create isogenic lines. An additional strong Htg QTL has been mapped on chromosome 9 (Htg9.1) in a second RIL population derived from L. sativa accession PI251246 and cultivar Salinas. Fine mapping is in progress to identify candidate genes associated with this QTL.

    A new project has been initiated to identify genes associated with the phenotypic plasticity of lettuce seed dormancy and germination characteristics in relation to the seed maturation environment. Seeds of the parental lines of the RIL populations described above were produced in low (25 ºC) and high (35 C) temperature greenhouses. The seeds produced in the two environments exhibited distinctly different phenotypic germination responses to temperature and light. We will attempt to map the loci associated with this plasticity in the RIL populations derived from these parents and establish whether it is associated with epigenetic DNA markers (methylation). Foundation seed from the two RIL populations are being produced to enable their production in multiple environments to facilitate this mapping approach.

    Down regulating expression of thioredoxin in wheat improved certain starch paste traits of both wheat cultivars. Certain farinograph and extensograph properties of the high gluten variety did not change, while farinograph properties of the low gluten variety decreased slightly compared to the parental control. Thus achieving PHS tolerance through modulation of thioredoxin expression resulted in higher yields but did not come at the expense of dough quality.

    We devised two approaches to extract and identify proteins undigested by pepsin in uncooked and cooked meal from numerous sorghum and corn varieties. The first approach involves extracting residues undigested after 2 h with Borate-SDS-ME and separated by PAGE. Improved separation of undigested sorghum proteins was achieved using NuPAGE Bis-Tris gels. Western blots, with antibodies against particular zeins and kafirins, were used to monitor fates of different kafirins and revealed differential digestion rates. A second approach involved extracting undigested residues sequentially with 60% t-butanol, 60% t-butanol-ME and Borate-SDS-ME. The second approach, also coupled with western analysis, revealed the following. (i) Existence of oligomeric forms of certain kafirins that differ in the degree of their susceptibility to pepsin digestion. (ii) Effect of cooking on the formation and digestion of the oligomers. (iii) Cross-linked forms of most kafirins became more resistant to digestion after cooking. (iv) Most alpha-kafirins are preferentially extracted in 60% t-butanol-ME while most gamma-kafirins are extracted in Borate-SDS-ME buffer. (v) Monomeric kafirins are resistant to pepsin digestion. (vi) Gamma-kafirins form a series of oligomers that exhibit differential resistance to digestion. Our results suggest that this presently described systematic approach to analyzing the digestion by pepsin of sorghum prolamins should lead to greater insights into the digestion of specific types of sorghum grain proteins.

    Comparative proteomic analysis of recalcitrant and orthodox Spartina seeds, using two-dimensional electrophoresis coupled to mass spectrometry, is being conducted to identify proteins associated with desiccation tolerance, as well as during cold stratification to determine protein changes that occur during loss of dormancy.

    The breeding system of sugar beet does not facilitate traditional genetic approaches to gene discovery, so populations were constructed using self-fertility as a means to genetically dissect seedling vigor traits. A Rhizoctonia seedling disease nursery was continued, and results demonstrated resistance to Rhizoctonia damping-off is present in sugar beet, and resistance has been incorporated into the newly released enhanced germplasms SR98 (PI 655951) and SR98/2 (PI 659754). A candidate gene transcript profiling was untaken to examine early growth stages during germination. Genes involved in stress responses, signaling and growth were activated within one hour after imbibition.

    To see whether TERF1 up-regulates PR- and cell wall-associated genes in seeds, Arabidopsis transgenic plants, in which TERF is chemically inducible, have been generated using the GeneSwitch technology (see below). Twenty-four independent, transgenic lines, which are resistant to hygromycin, have been isolated. T-2 seeds have already been produced. The progeny plants are currently subjected to genotyping and gene expression analysis to examine responses to the chemical inducer, methoxyfenozide. Arabidopsis siliques on the maternal plants were treated with the chemical inducer to cause NCED6 induction in developing seeds. Comparison of NCED6 expression between ligand-treated and -untreated siliques in the same flowering stem indicated that NCED6 was specifically induced by the ligand application. Wild-type siliques and untreated transgenic siliques also showed NCED6 mRNA signals, however they do not indicate leaky expression but reflect the native expression of NCED6 during the maturation stages. This established system allowed us to plan on and propose more experiments to analyze molecular changes downstream, which has led to a successful funding from American Seed Research Foundation.

    The role of polycomb complex in the regulation of rice endosperm development and grain filling was examined in rice. We purified the rice OsFIE2-polycomb complex using tandem affinity purification and demonstrated its specific H3 methyltranferase activity. We found that the OsFIE2 gene product was responsible for H3K27me3 production specifically in vivo and the gene expression was not regulated by imprinting. Genetic studies showed that a severe reduction of OsFIE2 expression led to completely endosperm-free seeds and a moderated reduction of OsFIE2 expression resulted in smaller seeds and loss of seed dormancy. Genome wide ChIP-seq analyses found that a large number of endosperm specific regulatory genes and storage nutrient metabolic pathway genes were directly regulated by H3K27me3 modification in the rice endosperm.

    In a previous study, we found that high temperature germination sensitivity was genetically controlled by a major QTL on the short arm of chromosome 3A (Qhtgs.osu-3A) in two winter wheat populations of recombinant inbred lines (RILs) generated from the locally adapted cultivars, 'Intrada' x 'Cimarron' and 'Jagger' x '2174'. In the present study, we tested the same set RILs of the Jagger x 2174 population for response in germination to low temperature, high temperature, and normal temperature. Results showed that Qhtgs.osu-3A disappeared when the seeds of the population were treated at 4 C for 3-7 days or were kept at normal temperature at 25 C. We have also developed markers for three genes, ABC-3A, HSP16, and MFT-3A, and mapped them on the distal end of the short arm of chromosome 3A. The MFT1 was associated with Qhtgs.osu-3A.

    The hormone ABA triggers wheat grain dormancy. The problem was that wheat varieties sometimes lack seed dormancy leading to a tendency for mature grain to germinate on the plant before harvest. This preharvest sprouting causes financial hardship for farmers because the grain must be used for feed. A technique was developed to identify ABA hypersensitive mutations in wheat resulting in increased grain dormancy which should provide increased resistance to preharvest sprouting. The increase in ABA-hormone response caused grain to loose dormancy more slowly after maturity, but did not eliminate the capacity to loose dormancy by after-ripening. GA signaling controls seed germination and plant height. The problem is to understand the mechanisms by which the GA stimulates seed germination and stem elongation. Research showed that GA stimulates seed germination and plant growth by causing two F-box proteins, SLEEPY1 and SNEEZY, to target master negative regulators called DELLA for protein destruction. SNEEZY destroys the subset of DELLA proteins that control plant height, but not the DELLAs that control seed germination. ABA insensitive mutants of hard red spring Scarlet wheat showed decreased seed dormancy and rapid after-ripening. Scarlet ABA-insensitive lines show initial seed dormancy levels and preharvest sprouting tolerance comparable to those of the original wild-type Scarlet. However, ABA-insensitive lines after-ripened more rapidly requiring only 3 weeks rather than 4 months of after-ripening to germinate.

    New work is underway to define the occurrence of the respiratory transition during maize seedling growth and development. We have begun to characterize germination and seedling growth protocols in the laboratory under light and dark conditions with the goal of comparing temperatures to ascertain what happens to the respiratory transition during germination and seedling growth at low temperatures. Currently we are evaluating seed lots. The respiratory transition has been studied at temperatures around 22-25 C but we do not know of studies investigating the impact of low temperature stress on these parameters. Two research projects have been initiated to answer a pressing question for the seed industry: "Can soybean seeds maintain viability and vigor in storage after 1 year?" The farmers' demand for treated soybean seed has grown from less than 8% treated soybeans in 1996, to more than 30% in 2008 (Munkvold, 2009), and all indications are that the trend is continuing. Soybean seeds exposed to inadequate storage environments in dealers' warehouses and farmers' storage sheds can lose seed viability and vigor. In the past, seed companies sold the returned untreated seed from dealer's warehouses in the commodity market. Today, seed companies must incur high cost in disposing of treated soybean seed in an environmentally safe way. Prolonging soybean seed viability and vigor in storage could save seed companies the cost of seed disposal.

    Objective 2. Determine and model the biotic and abiotic factors affecting seed germination, seedling emergence, and establishment of sustainable populations in natural and agro-ecological systems.

    Relationships were developed with seed producers in the emerging wildflower seed industry of the southeast US and restoration/conservation practitioners to identify key areas of seed biology research. This is evidenced by $180,336 in funded grants spanning 2009-12. Projects revolved around developing models to describe desiccation sensitivity in palms, germination physiology of orchids, genetic diversity among seed producing populations of a key species used in wild land restoration, cryopreservation of dune-stabilizing plant germplasm, and assessing the wildflower seed industry in Florida.

    Obligate root parasitic plants of the Orobanchaceae do not germinate unless they chemically detect a host plant nearby. Some members of this family, like Orobanche, are noxious weeds that cause heavy crop damage to agriculture worldwide. The endothelium, which is the inner layer of the testa, rapidly absorbs water. Its interconnected cells contain labyrinthine walls and filled with mucilage, facilitating water accumulation for germination that starts after receiving germination stimuli. Swelling of the endothelium also leads to the opening of a water gap at the micropyle. The perisperm cells underneath this gap mediate between the rhizosphere and the embryo, and are the likely location for the receptors of germination stimuli. The other perisperm cells, as also the endosperm cells and parts of the embryo, are loaded with lipids and protein bodies. Many plasmodesmata connect the perisperm cells to each other, and the cells near the micropyle tightly surround the emerging seedling. These perisperm cells, and also the proximal embryo cells, resemble each other in their cytoplasmic contents, and are actively involved in transfer of reserve nutrients to the developing seedling during germination.

    We are collaborating in a workgroup on Germination, Trait Coevolution, and Niche Limits in Changing Environments sponsored by the National Evolutionary Synthesis Center (NESCent) at Duke University. Though this group, we are working with colleagues at the University of Arizona (Larry Venable) and at the Institute of Botany, Chinese Academy of Sciences (Zhenying Huang) to conduct hydrothermal time analyses of seed germination of 13 native or introduced species. We applied hydrothermal time analysis to seed germination responses in relation to after-ripening to assess how changes in dormancy relate to their ecological fitness and demographic success. A manuscript is in preparation describing this work, as is a review of applications of hydrothermal time analysis to seed ecology. Hydrothermal time analysis was also applied to characterize germination and dormancy properties of herbicide-susceptible and herbicide-resistant populations of Echinochloa phyllopogon in California.

    Applications of abscisic acid (ABA) have been investigated as a method to enhance drought tolerance and condition transplants to better withstand post-transplanting field stresses. We are now developing foliar application methods with ABA to improve transplant quality while suppressing stem elongation rates in vegetable transplants grown in the nursery. We examined ABA foliar applications on transplant growth and yield of jalapeno pepper and triploid watermelon. In pepper, ABA was applied once (3.8 mM) at 22, 15, or 8 days before transplanting (DBT) or twice (1.9 mM) at 22 and 15 DBT. In watermelon, ABA was applied in the similar timings but with half the rate used for pepper. ABA was also evaluated as a means to control growth of mature triploid watermelon transplants (5-, 6-, 7- and 8-week old). In pepper, plant height showed a similar reduction of 10 to 14% for all ABA treatments. In contrast, reductions in shoot dry weight varied from 17 to 35%, with the strongest inhibition induced by the single application at 22 DBT. In watermelon, ABA did not affect plant height, but reduced shoot dry weight by 16 to 23%. In both species, these differences diminished gradually after transplanting and no yield difference was observed.

    Objective 3. Develop, evaluate, and transfer technologies to assess and improve seed and seedling quality, health, performance, utilization, and preservation.

    Carbon nano-tubes (CNTs) and nanocrystals were characterized and then applied to seeds as a dry power (CNTs only) or dispersed in water/agar (both CNTs and nanocrystals). Research was initially conducted by applying CNTs to 3 crop seeds: tomato, cucumber and soybean. This crop selection provides one of each of three different permeability types, permeable, selective-permeable and non-permeable, in our lab's model system. We observed a germination enhancement in all seed types even at the lowest concentration of CNTs. We saw improved conditions on plates and in roll towels. This may be due to disinfecting/filtering properties of CNTs or their ability to eradicate pathogens and/or enhance germination. Next, we tested 4 crop seeds (by adding switchgrass) with nanocrystals. We sought to determine if this nano-size material that is smaller than CNTs could penetrate seed coats. We conducted a survey of all four crop seeds to determine if the 10 nm nanocrystals would penetrate the seed coat during imbibition.

    We are making more intensive use of our X-ray analysis capabilities to develop protocols and information about the quality of our seed accessions beyond germination tests (standard protocol for germplasm centers). We've been collaborating with Dr. Mark Roh of the USDA-ARS in evaluating seeds of Cornus and Corylopsis by X-ray imaging. A visiting scholar (V. Neumann-Silva) from USP-ESALQ (Brazil) did a careful analysis of seed free space and vigor in Portulaca. A manuscript based on this research is in preparation.

    Bacterial Fruit Blotch (BFB) in cucurbits is a disease caused by Acidovorax avenae affecting worldwide watermelon and melon production. Currently there is a zero tolerance policy for BFB in growing facilities; however, there are no assays sensitive enough to reliably detect BFB in seeds. Two of the possible options existing for detection of BFB in early stage development of infected plants are Fourier Transform Infrared Spectroscopy (FTIR) and the use of chemosensory analysis. FTIR analysis was conducted in dry seeds, imbibed seeds, germinating plants, and small seedlings in order to determine the limit of detection for the sensors. Measured spectrums and olfactory profiles will then be analyzed for differences. Early trials have demonstrated the feasibility of using FTIR to rapidly and nondestructively differentiate contaminated seeds from non-infected seeds. Also FTIR has nondestructively differentiated alive and dead seeds from within a seed lot.

    Recent human disease outbreaks of bacterial pathogens transmitted on vegetables, especially fresh spinach, have raised questions about the potential for seed transmission of human pathogens. The bacterial populations of spinach seed and seedling leaves were compared using DGGE, to assess bacterial community richness, and real-time PCR to compare the abundance of select phyla (total bacteria, Actinobacteria, Bacteroidetes, Firmicutes, a-Proteobacteria and b- Proteobacteria). To determine the effect of environment, the plants were grown in field and growth chambers. Vertical transmission of bacterial community members was evident because the developmental stage of the plant affected the richness and abundance of select bacterial phyla. The bacterial richness of plants grown in the two environments was not affected. However, a greater number of bacteria were associated with field grown samples in comparison to those produced in growth chambers. A statistically significant interaction existed between growth stage and environment for each of the selected phyla. Populations on cotyledons were smaller than mature leaves, but were not significantly different than the 3-4 leaf stage seedlings. The culturable populations of bacteria on seeds (~5 log CFU/g) were significantly smaller than determined using real time PCR (~7 log copies). Vertical transmission of bacteria from seeds to seedlings, suggests that seed may have evolved to ensure the transmission of certain bacteria.

    Switchgrass (Panicum virgatum L.) is used horticulturally as an ornamental and agronomically as an animal feedstock and a putative bio-energy crop. Genetic transformation, using somatic embryogenic (SE) callus derived from mature seeds, is one strategy for improving switchgrass traits. A superior switchgrass line, HR8, was developed in using recurrent tissue culture selection from cv. 'Alamo' that has low dormancy and produces a high percentage of somatic embryos. Eighty two percent of HR8 seeds germinated after harvest comparing to 26.8% for unselected 'Alamo'. HR8 seeds that germinated produced 84.9% SE callus.

    The respiratory activity of individual seeds during imbibition and germination is being assessed using the Q2 instrument from ASTEC Global. Initial seed respiration during imbibition and germination is being characterized in relation the effects of respiratory inhibitors to identify the metabolic pathways underlying different components of the oxygen uptake patterns exhibited by imbibed and germinating seeds. New methods were developed to analyze Q2 data that makes it amenable to use in population-based threshold models. We are using this approach to analyze and model respiratory responses to inhibitors, water potential, temperature and aging.

    The effects of seed drying methods on seed longevity are being assessed. Seeds exhibit hysteresis in their moisture content/relative humidity relationships, or isotherms, depending upon whether they are losing or absorbing water. Seeds that are drying have moisture contents that are generally 0.5 to 1% higher in moisture content than seeds that are hydrating to the same equilibrium RH from a lower moisture content.

    A second year of studies has confirmed that this difference in seed moisture content results in corresponding differences in seed longevity. Based on these results, simple modifications to seed drying procedures may be able to extend seed longevity in storage. With funding from the USAID-funded Horticulture Cooperative Research Support Program (HortCRSP), we are demonstrating and disseminating desiccant drying bead technology for seed drying and storage by smallholder farmers in India, Nepal, Bangladesh, Kenya, and Tanzania. Seed drying beads can dry seeds efficiently in hermetic containers, and can be regenerated by heat and reused indefinitely. This technology has considerable promise for improving seed quality and longevity in humid regions.

    Impact Statements:
    1. University of Florida - The impact of our research is that restoration practitioners and seedling producers can better plan and coordinate in terms of seeding activities for restoration of degraded lands. Similarly, conservation practitioners can adapt methods for ex situ activities. We also can develop targeted research programs to enhance the wildflower seed industry.
    2. Cornell University - Our tests do not support the idea that the CNTs penetrate the seed coat of tested seeds. Crystals readily penetrated seedcoats of soybean, tomato with very few in switchgrass. Nanocrystals were observed in cotyledons of soybean and embryo/endosperm of tomato. In cucumber crystals appeared in the cotyledon envelope. Nanocrystals act as a fluorescent marker for initial water uptake.
    3. Ohio State University - Non-deep physiological dormancy appears dominant in Phlox; with variations among accessions. Seeds of P. paniculata produced in the green house appear to have different dormancy characteristics than the progenitor seeds from wild populations. Large variations exist in the response of Rudbeckia fulgida seeds to GA and to combinations of GA and cold treatments. Rapid systems to assess vegetable and other crop seed quality and vigor have been refined.
    4. Virginia Tech - FTIR can be used to nondestructively differentiate between clean and infected seeds. Switchgrass line HR8 has low seed dormancy and produces a high percentage of somatic embryos from mature seeds for genetic transformation. Supplied thermogradient tables to aid research in the seed industry. Orobanchaceae seeds are successful parasites in part due to a unique seed anatomy that allows seeds to hydrate quickly and sense chemical signals from suitable hosts in close proximity.
    5. South Dakota State University - Map-based cloning of the SD7-1 seed dormancy QTL answered the long standing question that the association between resistance to pre-harvest sprouting and red grain color in cereal crops arises from pleiotropy and the dormancy gene cannot be used to improve white grained colored cultivars for resistance to pre-harvest sprouting by conventional breeding approaches.
    6. University of California-Davis - Seed quality analysis protocols based upon respiratory patterns of germinating seeds are being developed and distributed. Desiccant-based seed drying methods can provide a simple, economical method to dry seeds in humid environments. Simple modifications of seed drying protocols could significantly improve seed longevity in storage. Genetic capacity for high temperature germination would improve germination and stand establishment at high temperatures.
    7. University of California-Berkeley - Changes in the expression of different forms of the regulatory protein, thioredoxin, can improve nutritional and agronomic properties of wheat. The work also opens a new door to our understanding of redox regulation and its role in seed development and germination.
    8. Louisiana State University - Expression patterns suggest that prevention of programmed cell death and protein stabilization by LEAs (late embryogenesis abundant proteins) during seed drying, as well as detoxification of reactive oxygen species during subsequent germination are important for viability of drying-tolerant seeds. These data have identified putative targets for improving seed stability of Spartina alterniflora.
    9. USDA-ARS and Michigan State University - The beet gene knowledge base is a unique and vital research tool to assess biochemical and genetic responses to stress during germination, and now allows scientists a better understanding and predictive ability to improve seedling vigor required for sugar beet to germinate and be productive in all environments.
    10. Oregon State University - TERF1-inducible Arabidopsis seeds will enable finding the mechanisms of induction of PR- and cell wall-associated genes. TERF1 induction will also provide an excellent opportunity for technology development for seed germination control. The GeneSwitch approach can be used to prevent pre-harvest sprouting problems in cereal seeds such as wheat and barley.
    11. Mississippi State University - Our study demonstrated that epigenetic regulation is essential for rice endosperm development and probably play a critical role in seed dormancy regulation. Our results suggest that understanding epigenetic regulation is critical for the improvement of seed yield and storage nutrition quality. In addition, our comparative studies of the four ChIP-Seq analytical algorithms provide a guide line for future ChIP-Seq data analysis.
    12. Texas AgriLife Research, Texas A&M System - ABA foliar application is effective in size control of vegetable transplants and its growth inhibiting effects are transient. In triploid watermelon, a single foliar application of ABA was more effective on improving growth and yield when applied to 5-week old transplants as compared to untreated 5-week old or ABA-treated older transplants (6 to 8 weeks).
    13. Oklahoma State University - Results demonstrated that the effect of Qhtgs.osu-3A on seed germination was regulated by temperature.
    14. Washington State University - New wheat lines can be used to breed for resistance to preharvest sprouting. Two F-box proteins could be altered to make shorter plants that are more resistant lodging without causing poor germination and seedling emergence. Allelles from Scarlet ABA-insensitive lines could be used to improve seedling emergence of winter wheat varieties.
    15. Iowa State University - The findings from our research are important to crop-protection chemical companies, because seed treatments are one of the fastest growing segments of the crop protection market. Chemical companies would benefit from better storage methods for treated soybean seed. The respiratory transition should be an important element of genotypic variation for low temperature stress tolerance. As production costs rise, it becomes more important to achieve target populations.
    Last Modified: 01-May-2012

    Date of Annual Report: 06/11/2013

    Report Information:
  • Annual Meeting Dates: 04/25/13 to 04/26/13
  • Period the Report Covers: 10/2012 to 09/2013

  • Participants:
    Brief Summary of Minutes of Annual Meeting:
    April 25 8:30 a.m. Susana Goggi: calls meeting to order, presents opening remarks, requests participants introduction, and requests President Xingyou Gu to chair the meeting.

    8:40 a.m. Xingyou Gu: 1) Approval of 2012 Minutes (Kent Bradford motion, Hector Perez 2nd, and all in favor). 2) Hector Perez (the past president) provided a brief report on the state of W-2168 membership. New possible members were discussed. 3) Leadership structure: Kent leads the Secretary-elect Nominating Committee; current president, Xingyou Gu; president-elect, Susana Goggi; Secretary, Pablo Jourdan. 4) Discussion of possible seed biology symposium: after review of W-3168 proposal and further discussion on sites and other seed symposia currently on the schedule for the next few years, no actions were taken. 5) International Innovation Report from Research Media Ltd. was discussed and a decision was made not to take action. 6) Manjit Misra welcomed the group. 7) Donn Thill, Administrative Advisor, joined the meeting via phone link: he complemented the group regarding the proposal re-write and discussed next steps in the review and renewal process. 8) Hans Arne Jensen shared his views regarding the status of ISTA laboratories and Seed Testing in Europe;

    11:00 a.m. State reports started: Dr. Alan Taylor and Dr. Kent Bradford presented their annual reports. Questions and discussions were made after each of the report.

    2:00 p.m. State reports continued: Dr. Robert Geneve and Dr. Greg Welbaum presented their annual reports. Questions and discussions were made after each of the reports.

    3:00 p.m.: Tour of Seed Science Center at Iowa State University.

    April 26 8:30 a.m.: Xingyou calls meeting to order. Welcome by Dr. Joe Colletti, Senior Associate Dean, ISU College of Agriculture and Life Sciences, and Associate Director of the Experiment Station.

    9:00 a.m. State reports continued: Drs. Susana Goggi, Hector Pérez, Dr. Pablo Jourdan, Bruce Downie, Sabry Elias, and Xingyou Gu presented their annual reports. Questions and discussions were made after each of the reports.

    11:30 am.: Reports of Secretary-elect, site location meeting for 2014 annual meeting, and other new business. Bob Geneve was named as the incoming Secretary. Meeting site for 2014 annual meeting will be at Ohio State University in July.

    12:15 pm: Adjourn.

    Objective 1. Identify and characterize biophysical, biochemical, genetic, and environmental factors regulating or influencing seed development, germination, vigor and dormancy

    Bradfords group (CA) cloned a QTL associated with high temperature germination (htg) in lettuce, which was identified as the gene LsNCED4 encoding the enzyme 9-cis-epoxycarotenoid dioxygenase 4 in the abscisic acid (ABA) biosynthetic pathway. An additional htg QTL was mapped on chromosome 9 (Htg9.1). Fine mapping of the new QTL is in progress to identify candidate genes. An atlas of gene expression during imbibition and germination of four genotypes of lettuce under different temperature and ABA conditions is being developed using RNA sequencing (RNASeq). The seed dormancy gene DELAY OF GERMINATION 1 (DOG1) in Arabidopsis has been determined to be involved in high temperature germination and the DOG1 homologs have been isolated from lettuce to characterize their expression and function(s) in lettuce seed dormancy.

    Pérezs group (FL) focused on seed development, dormancy, and germination ecology of wildflowers and some warm-season grasses. Seed development studies investigated desiccation tolerance, germination ability, and aging stresses in developing seeds and embryos. Seed dormancy studies addressed dormancy developmental controls, dormancy mechanisms expressed by seeds, and methods to alleviate these mechanisms. Germination ecology research revolved around germination timing and the seeds interaction with the environment to alleviate dormancy and promote germination.

    Nonogakis group (OR) performed research on mechanisms of tissue specific gene expression in tomato seeds. GeneChip analysis was used to identify multiple genes whose expression was enriched in the micropylar region of endosperm (endosperm cap) in tomato seeds. Some of these genes contained the conserved DNA motifs, suggesting their regulation by a common regulatory protein. The TOMATO ETHYLENE RESPONSE FACTOR1 (TERF1) was identified as a potential regulatory protein that enhances expression of multiple endosperm cap-specific genes. Function of hormone metabolism genes in Arabidopsis has been characterized.

    Gus group (SD) continued the research on seed dormancy and also initiated a project on seed longevity in rice. The project of cloning the qSD12 QTL was focused on developing isogenic lines for the natural alleles of three candidate genes and transgenic T2-T4 lines with the candidates. Haplotypic analysis for the qSD12 region was conducted for 400 lines of wild, weedy and cultivated rice. Genetic and physiological analyses were conducted for two chromosomal segments each containing the QTL clusters qSD1-2/qPH1 and qSD7-2/qPH7 for seed dormancy/plant height. QTL and haplotype analyses for seed dormancy and its related adaptive traits were conducted for U.S. weedy rice populations. Seed longevity was assessed for a segregating population of seeds aged under the room condition for about 10 years. A new population of >400 recombinant inbred lines derived from a line of U.S. weedy rice was advanced to the F8 generation to investigate ecological and genetic mechanisms of seed longevity under controlled and field conditions.

    Baskins group (KY) investigated mechanisms of breaking seed dormancy in various species including Solanum rostratum, Cardiocrinum giganteum var. giganteum, Alyssum minus, Astazgalus arpilobus and Suaeda corniculata, gave eight lectures on seed dormancy and seed bank, and published 18 papers with various collaborators.

    Downies group (KY) characterized the role of CRYPTOCHROME 1 and 2 in Cryptochome control of blue light induced dormancy in barley and assessed the function of the ATP-dependent, DEAD-box, RNA-Helicase PRH75 in seed viability and morphology.

    Geneves group (KY) worked on Eastern gamagrass, a warm-season, perennial grass native to large areas across North America. Plantings are commonly established from seed, but seed quality is a barrier to establishment. Standard laboratory purity methods and germination testing protocol needs to be standardized for routine assessment of eastern gamagrass seed lots for planting purposes. A survey of 21 labs determined that at least 9 germination procedures are currently in use. A comparison of a single seed lot as reported by 10 labs was used to determine variation in test results. Additional seed lots were germinated on substrates with and without potassium nitrate (KNO3) and subjected to prechilling and presoaking treatments. The portion of the seed quality and assessment portion of the gamagrass work has been completed. Work is now beginning of the treatments for improved dormancy release and increased first year stand.

    Stebers group (WA) worked on the wheat preharvest sprouting problem and mechanisms of the plant hormone GA in seedling growth and development. ABA-insensitive mutants in the genetic background of the dormant red wheat cultivar Scarlet were characterized.

    Cohns group (LA) continued working on physiological/molecular mechanisms of seed dormancy, seed desiccation intolerance (recalcitrance) and related problems in the marsh grass. Spartina alterniflora is an important component for successful maintenance and remediation of Louisiana coastal stabilization. Differentially expressed proteins between the dormant and nondormant state of Spartina alterniflora and Spartina pectinata seeds were isolated by two-dimensional gel electrophoresis comparisons and sequenced using mass spectrometry. The effects of drying rate and drying temperature on recalcitrant seed death will be re-examined.

    Jangs lab (OH) has been looking at the involvement of tandem CCCH zinc finger proteins (TZFs) in hormone-mediated developmental and environmental responses. Several TZFs were characterized for expression profile during seed germination in Arabidopsis and their responses to the ABA and GA hormones.

    Mercers lab (OH) has been exploring the germination characteristics of different landraces of maize from southern Mexico to determine how each might respond to potentially altered climatic conditions.

    Jourdans lab (OH) at the Ornamental Plant Germplasm Center (OPGC) continued several projects. One project was to study the biology of Begonia seeds and to develop an appropriate germplasm collection that emphasizes seed storage. Efforts have made to develop rapid and efficient quantitative seed germination tests for the minute seeds; assess basic biochemical properties (carbohydrate, oil, protein content) and desiccation tolerance (seeds of some species appear to be orthodox and relatively long-lived), and develop accelerated aging protocols for evaluation of vigor and prediction of long-term storage. The other project was to develop efficient seed production protocols for the 200 accessions of Phlox that have been accumulated and maintained as plant samples over the past two years. Controlled pollination techniques and seed dormancy characteristics in P. paniculata were investigated. The OPGC has also been exploring the germination patterns of different species of Rudbeckia.

    Obendorfs lab (NY) conducted research on several projects to determine if changes in soluble carbohydrates in hypocotyl and radicle tissues correlate with differential tissue survival after drying seedlings in buckwheat (Fagopyrum esculentum Moench) embryos, if raffinose and stachyose are unloaded from seed coat to developing embryos in soybean, and whether increasing the supply of free cyclitols to immature embryos increases accumulation of galactosyl cyclitols in soybean lines selected for low raffinose and low stachyose (LRS) or low raffinose, low stachyose and low phytin (LRSP1, LRSP2) concentrations in mature seeds, as compared with a CHECK line with normal raffinose, stachyose and phytin.

    Objective 2. Determine and model the biotic and abiotic factors affecting seed germination, seedling emergence, and establishment of sustainable populations in natural and agro-ecological systems

    Bradfords group (CA) collected accessions of Brachypodium hybrida, which is a moderately invasive, introduced grass species in California, and all were determined to be polyploid, although both diploid and tetraploid forms are present in its native range in Turkey. The collections from the state and the accessions from Turkey and other locations are being used to study the inherent, variation or plasticity in flowering time and seed dormancy to test the hypothesis that polyploids may have greater genetic variability in the flowering and seed dormancy traits, which may explain the preponderance of polyploids among invasive species.

    Pérezs group (FL) developed models to describe desiccation sensitivity in palms, cryopreservation of dune-stabilizing plant germplasm, examined tolerance of U. paniculata seeds to various concentrations of a commercial biocide, evaluated the chronic and range-wide loss of seed viability loss in Aristida stricta, and assessed cryo-storage tolerance of 4 U. panicualta seed accessions.

    Baskins group (KY) investigated dormancy cycling in the cold desert ecosystem and soil seed banks in forests of northern Iran, and helped pull together the literature on the ecological importance of mucilage in seed germination ecology.

    Downies group (KY) assessed the homologs of Arabidopsis and soybean LATE EMBRYOGENESIS ABUNDANT (LEA) protein client proteins, identified preferred LEA binding partners, characterized three SEED MATURATION PROTEIN1 (SMP1) binding proteins for functions in inducing thermodormancy in Arabidopsis, and constructed a web page as a data repository as part of a project examining Arabidopsis and soybean LEA protein interacting client proteins.

    McGraths group (MI) has focused on stress responses of seed germination and seedling field emergence/persistence in sugar beets to develop healthy, vigorous, and productive field populations of the crop. Despite planting high-quality, technically-augmented seed for growers with very high germination (>92%), field emergence and persistence continues to hover at ~60% in Michigan. Research has generated four transcriptome datasets of a high vigor variety germinated in stressful environments, including water, hydrogen peroxide, sodium chloride, and hydrogen peroxide plus sodium chloride. From over 200 million sequences assessed, and in comparison to water, each treatment showed between 75 and 100 statistically significant, differentially expressed genes, with approximately 10% overlap between any pair of treatments, and only seven shared among treatment comparisons. Approximately 50% of these have no descriptors yet found in nucleotide sequence databases, suggesting germination genes are less well represented in comparative genomics resources.

    Welbaums group (VA) worked on Bacteria Fruit Blotch (BFB), a seed-transmitted disease of cucurbits caused by Acidovorax avenae subsp citrulli (Aac) that is one of the most devastating diseases that affects both watermelon (Citrullus lanatus) and cantaloupe (Cucumis melo) production worldwide. The genome of Aac strain AAC00-1 (group II) was sequenced. The cucurbit germplasm from the USDA collect was screened to identify genetic resistance to different type III effectors. The group was also involved in a project to assess the potential for biological contamination on greenhouse lettuce and basil plants grown in aquaponic wastewater.

    Objective 3. Develop, evaluate, and transfer technologies to assess and improve seed and seedling quality, health, performance, utilization, and preservation.

    Bradford group (CA) has developed new methods to analyse Q2 data to monitor the respiratory activity of individual seeds during imbibition and germination, which makes it amenable to use in population-based threshold models. Effects of respiratory inhibitors on seed oxygen consumption during imbibition and germination have been characterized. Research has been started to identify metabolites and metabolic pathways underlying different components of the oxygen uptake patterns exhibited by imbibed and germinating seeds. The effects of seed drying methods on seed longevity are being assessed. The Q2 instrument has been utilized to show that seed respiration rates are highly correlated with germination rates (timing). Semi-automated respiratory assays in the Q2 can be used to collect germination rate data without requiring labor-intensive repeated observations of germination tests. With funding from the USAID-funded Horticulture Cooperative Research Support Program (HortCRSP), a desiccant drying bead technology was developed for seed drying and storage by smallholder farmers in India, Nepal, Bangladesh, Kenya, and Tanzania. The knowledge and techniques developed by this group were presented in five seminars, in addition to publications.

    Nonogakis group (OR) developed a gene switch system for the Arabidopsis NCED gene, which provided a proof of concept for modification of seed performance by hormone metabolism engineering. While the Gene Switch system is useful for agriculture, chemical application may not be practical for large-scale agriculture. Therefore, a novel system that could cause spontaneous seed dormancy that does not require chemical induction has also been tested. Considering potential technology development as an outcome of this study, the concept has been tested using genes isolated from wheat and sorghum. The NCED gene, one of the rate-limiting genes for ABA biosynthesis, has been isolated from sorghum and fused with the promoter sequence of EARLY METHIONINE-LABELED, one of seed-specific and ABA-regulated genes, from wheat. Arabidopsis plants expressing this chimeric gene have been generated and tested for seed germination performance.

    Gus group (SD) continued the project to pyramid the dormancy-enhancing alleles at four QTL detected/reported in the previous research to improve the resistance to pre-harvest sprouting for local spring wheat cultivars with white grain color.

    Taylors group (NY) focused on mechanisms and technology development of breaking seed dormancy in switchgrass (Panicum virgatum). A major limitation of growing switchgrass is the poor and erratic stands of the crop during the first year due to seed lot quality and dormancy. A series of experiments were conducted to examine effects of the plant hormones/growth regulators GA, ABA, ACC and fluridone on germination capability. Research on seed priming using the varieties Kanlow and Blackwell was conducted to develop a technology to promote germination, especially under environmental stress.

    Welbaums group (VA) worked on the development of several germination techniques. Basil is a popular herb with little information about its base, ceiling and optimal germination temperatures. Six different basil genotypes were compared for responses to high, low, and optimum temperatures. Thermogradient tables were used to generate a wide range of temperatures for seed testing on a flat surface. New thermogradient tables were designed. A preliminary gradient table has been compared for germination performance of a wide range of peanut genotypes. FTIR (Fourier transform infrared spectroscopy) is a technique, which is used to obtain an infrared spectrum of absorption, emission, photoconductivity or Raman scattering of a solid, liquid or gas. An FTIR spectrometer simultaneously collects spectral data in a wide spectral range. A mathematical algorithm converts the raw data into the actual spectrum. This technology has been evaluated for its possible use for the rapid non-destructive assessment of seed and plant disease infection. The other activities included teaching a web-based, asynchronous distance learning Vegetable Seed Production Course and organizing the XXVIII International Horticultural Congress on Science and Horticulture for People (IHC2010) in Lisbon, Portugal.

    Goggis group (IA) continued seed physiology and pathology programs. The seed physiology program can be organized into two areas of research: how to select, breed, and produce soybean and corn cultivars with enhanced nutritional properties and other value-added traits and high seed quality; and how to minimize adverse effects of production environments on seed quality and storage life of soybean and corn. Current projects include genome fluidity and stress-induced changes in the soybean genome and their effect on seed and agronomic characteristics; seed storage in soybean and the effect of soil fertility and the use of seed treatment to prolong seed life. The seed pathology research program can be organized into three areas: integrated management of multiple pathogens and pests attacking seeds and seedlings; seed health test development; and infection of corn by mycotoxin-producing fungi. Current disease management projects include: investigations into various aspects of soybean seedling blight and root rot caused by Fusarium species; management of multiple maize and soybean seedling pathogens through seed treatment combinations; and the evaluation of seed treatment effects on maize head smut through a PCR-based seedling assay. The development of seed health test method has been focused on seed-borne diseases of edible beans (bacterial wilt and Fusarium wilt) and tomatoes (Potato spindle tuber viroid and other viroids). Mycotoxin-related projects include assessing the role of mycotoxins in seedling diseases of maize, soybean, and wheat; and evaluation of the impact of transgenic insect resistance on the quality of maize grains for ethanol production. Other activities include leadership of the distance-education Graduate Program in Seed Technology and Business, and seed health testing standardization efforts through the US National Seed Health System.

    Jourdan and Bennetts group (OH) worked on developing protocols and information about the quality of seed accessions in the Ornamental Plant Germplasm Center beyond germination tests (standard protocol for germplasm centers) by more extensively use of X-ray analysis capabilities.

    Lopes and Bennetts group (OH) assessed seed germination and vigor from different cultivars of snap beans (Phaseolus vulgaris L.) varying in seed coat color and seed size after being submitted to drum priming and subsequent drying treatments. SVIS and other seed quality tests (including seedling emergence in greenhouse and field studies) have been conducted.

    Leskovars group (TX) investigated effects of plant hormones on seedling establishment and quality. Exogenous abscisic acid (ABA) was applied to induce leaf chlorosis in various vegetable and ornamental crops, a potential problem for its commercial use. One mechanism generally proposed for ABA-induced chlorosis is senescence by stimulated ethylene production. ABA accumulation during water stress also inhibits leaf expansion to limit plant water loss. When this acclimation is induced by exogenous ABA, it is followed by rapid leaf expansion, with leaf area eventually recovering to the control level, as shown in the previous research. The group further investigated the effect of ABA on Arabidopsis leaves at different stages of development and expansion to determine if the hormone inhibits cell expansion and/or cell division. Research was also conducted to determine the relation between chlorophyll loss and ABA concentrations in muskmelon seedlings.

    Impact Statements:
    1. Introgression lines with the LsNCED4 alleles are being distributed to breeders and seed companies for integration into lettuce cultivars to alleviate thermoinhibition and improve efficiency of lettuce production in desert growing areas (CA).
    2. The methods developed based on relationships between seed moisture content and seed equilibrium relative humidity (RH) can be used to enable simple monitoring of seed storage conditions. Tools have been developed to easily convert seed equilibrium RH to seed moisture content and vice versa. Modifications of seed drying protocols could significantly improve seed longevity in storage (CA).
    3. The desiccant-based seed drying methods are being implemented in both large- and small-scale applications for drying seeds and other horticultural products. This technique provides a simple, economical method to dry seeds to safe moisture contents in humid environments without direct heat (CA).
    4. Respiratory patterns of imbibed/germinating seeds can be used to substitute for labor-intensive seed germination rate assays, enabling more efficient utilization of seed quality tests such as controlled deterioration (CA).
    5. The synthesized information on the response of wildflower seeds to simulated seasonal and constant temperatures could facilitate the production of pre-variety wildflower germplasm on both a regional and a global scale. With the information, restoration practitioners and seedling producers can better plan and coordinate in terms of seeding activities for restoration of degraded lands (FL).
    6. The research on palms linked the ability of embryos to accumulate dry matter throughout development with desiccation sensitivity and developed a model for allowable cell shrinkage. The model provides a framework to quantify desiccation tolerance via mechanical strain when embryo cells shrink during drying (FL).
    7. The information on the interactions between desiccation and freezing on the germination of Uniola paniculata, a dune-stabilizing plant essential to coastal restoration activities, could be translated into the development of effective ex situ storage and germination strategies (FL).
    8. The Gene Switch system developed by Nonogakis group allows hormone metabolism engineering in seeds, which can be applied to multiple genes associated with seed development and germination. This system opened a possibility to develop a robust technology to modify seed performance, and can be utilized to prevent pre-harvest sprouting in cereal crops (OR).
    9. The identification of the endosperm cap-specific genes in tomato revealed the involvement of ethylene signal transduction in the regulation of germination, which provided new knowledge in seed biology. The promoters of the endosperm cap-specific genes provide a new tool to drive tissue-specific gene expression in tomato seeds (OR).
    10. The isogenic lines for the naturally occurring alleles isolated from weedy rice at the qSD12 seed dormancy locus and the advanced transgenic lines developed with three different candidate genes of qSD12 are not only critical for the on-going project but also useful for breeders to manipulate cultivars for germination capability (SD).
    11. The research on the two clusters of QTL for both seed dormancy and plant height revealed that the development of these two traits share genetic and molecular pathways, and the semi-dwarf genes present in cultivars have some effects on resistance to pre-harvest sprouting, and the newly identified QTL allele from weedy rice could be used to modify plant height and germination capability in the crop (SD).
    12. The research in switchgrass revealed that the constant warm temperature induced secondary dormancy, GA treatments were only effective if a fluridone treatment was given, combinations of GA, ACC and fluridone resulted in the highest germination, but fluridone and GA could induce abnormal seedling growth and development. Priming increased germination, but the optimal effect varied with varieties and seed lots. A patent application was submitted on the controlled release of seed and soil treatments triggered by pH change of growing media (PCT/US2011/033420) (NY).
    13. Black seeds of the cold desert halophyte Suaeda corniculata buried in the field undergo a dormancy/nondormancy cycle and also seasonal cycles in their sensitivity to salinity (KY).
    14. Mucilage on seeds of the cold desert ephemeral Alyssum minus serves as a water reserve for germination when there is water stress (KY).
    15. Seeds of Cardiocrinum giganteum var. giganteum for the Himalayas have the potential to germinate under the climatic conditions of northern Japan, and thus there is a potential (at least from a seed germination perspective) that this beautiful lily could become invasive (KY).
    16. Soil seed bank of the old-growth forests in northern Iran do not contain seeds of climax trees, thus if forests are disturbed/destroyed the revegetation will be via secondary not primary forest species (KY).
    17. Seeds of Geranium carolinianum become sensitive (but not permeable) during summer, and when the temperature decreases to about 20 C in autumn they become permeable (KY).
    18. Cryptochome1, not Cry2, is responsible for the blue light enhancement of seed dormancy in barley. An ATP-dependent, DEAD-box RNA helicase has been shown to suffer loss of helicase function upon thermal insult and, in the short term (4 hours or less), which is reversible by incubation by PIMT and AdoMet. Loss of function alleles demonstrated that this helicase is essential for life (KY).
    19. LEA protein homologs of diverse species have specific target proteins with which they interact. Mutation of the LEA-like SEED MATURATION PROTEIN1 (At3g12960) resulted in seeds that were incapable of entering thermodormancy. This LEA and its soybean homolog GmPM28 (Glyma08g18400) repeatedly pulled down fragments of three proteins, identifying preferred LEA protein binding partners. Mutants of these protein-coding genes (AT1G30610, AT1G15290, and AT1G15280) failed to enter secondary dormancy (KY).
    20. Seed testing procedures have been standardized for testing gamagrass seed samples. For a routine laboratory standard germination test, seed analysts can use either blotter-lined Petri dishes or rolled towels as the substrate in the temperature regime of 20/30 °C with counting dates of 7 and 14 days (KY).
    21. ABA-insensitive mutants in dormant red wheat did not cause a decrease in initial seed dormancy or preharvest sprouting susceptibility, but did result in more rapid loss of dormancy with dry after-ripening. This rapid dormancy loss improved seed germination and emergence, and provides a genetic strategy to improve emergence of red wheat varieties (WA).
    22. Increased accumulation of the GA hormone receptor promoted the germination of dormant seeds, but the seedlings were underdeveloped and weak unless they were also exposed to GA hormone prior to seed germination. This suggests that increased GA signalling can be used to promote early growth, development, and vigor of germinating seedlings thereby alleviating problems with poor seedling emergence, a serious problem especially in semi-arid regions where seeds must be planted deeply to reach stored soil moisture (WA).
    23. Characterization of Aac strains revealed the existence of at least two well-differentiated groups of the disease. The first group includes strains that were mainly isolated from cantaloupe and pumpkin. While the second group represents the typical BFB strains that affect watermelon (VA).
    24. New thermogradient tables were designed and built for the seed industry through the company TASCO in Christiansburg, VA (VA).
    25. The knowledge generated by the seed physiology and pathology research programs in Iowa help seed industries and bioenergy companies to breed improved varieties, prolong the life of soybean seeds in storage, and facilitate the production of pathogen-free, high quality seed; and facilitate the movement of seeds worldwide (IA).
    26. Seventy-eight differentially expressed protein spots between the desiccation intolerant (DI) and tolerant (DT) Spartina genotypes were sequenced, and 68 of them have homologue matches in existing databases. The fate of these proteins was followed when DT seeds germinated and progressively lost desiccation tolerance as seedlings. All spots were present in small DT seedlings and vanished when the seedlings lost desiccation tolerance. These observations are consistent with a role for these proteins in seed desiccation tolerance and identified putative targets for improving seed stability of Spartina alterniflora (LA).
    27. Stress germination transcriptomes were obtained and found to contain many statistically significant differentially expressed genes for which no function can yet be deduced. Two biochemical pathways that influence seed germination and seedling vigor in ways that can improve emergence potential have been identified in sugar beets. Current results will be used to narrow the definition of seedling vigor to specific genes involved in germination under stress conditions (MI).
    28. Three tandem CCCH zinc finger proteins (TZFs) were identified specifically expressed in seeds and involved in abscisic acid- and gibberellic acid-mediated growth in Arabidopsis. The expression levels of these TZFs declined during seed imbibition, up-regulated by ABA and down-regulated by GA. Results of gene expression analysis indicate that these TZFs affect seed germination by controlling genes critical for ABA and GA responses (OH).
    29. Post-harvest methods were developed to enhance soybean, castor bean, snap bean, and tomato seed quality and protect seedlings from detrimental effects of sub-optimal growing conditions and pests/diseases (OH).
    30. Refined systems were developed to rapidly assess vegetable and other crop seed quality and vigor (OH).
    31. Epigenetic mechanisms mediated by polycomb complex are involved in the regulation of endosperm and embryo development and probably also play a critical role in the regulation of seed dormancy in rice. Candidate RNAs in the regulatory networks have been identified. The knowledge of epigenetic regulation will contribute to the improvement of seed yield and storage nutrition quality (MS).
    32. ABA-induced chlorosis was leaf age-dependent and can occur independently of ethylene. ABA inhibited leaf expansion by limiting cell expansion, not the number and size of stomata. Thus, ABA-induced inhibition of leaf expansion is a mechanism to conserve water by limiting increases in non-stomatal evaporative area, as opposed to stomatal closure that reduces transpiration. This mechanism may not limit plant growth and photosynthetic capacity, as leaves maintain both cell division and stomatal formation (TX).
    33. The research on common buckwheat suggested that an apparent shift in biosynthetic pathways within hypocotyls resulted in raffinose and stachyose accumulation, but not fagopyritols, during drying of seedlings. The survival of hypocotyl tissues correlated with an increase in concentrations of raffinose, stachyose, and sucrose (NY).
    34. During soybean seed development, seed coat tissues accumulate raffinose and stachyose. Traces of raffinose can be detected in cotyledons of young developing seeds and infrequently in seed coat cup exudates at mid-seed fill. The biochemical research provided evidence that soybean seed coat unloaded raffinose in very small amounts, which may explain the presence of trace amounts of raffinose in embryo tissues of young seeds (NY).
    Last Modified: 19-Jun-2013
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