Development of SSR markers and analysis of diversity in Turkish populations of Brachypodium distachyon

Background: Brachypodium distachyon (Brachypodium) is rapidly emerging as a powerful model system to facilitate research aimed at improving grass crops for grain, forage and energy production. To characterize the natural diversity of Brachypodium and provide a valuable new tool to the growing list of resources available to Brachypodium researchers, we created and characterized a large, diverse collection of inbred lines. Results: We developed 84 inbred lines from eight locations in Turkey. To enable genotypic characterization of this collection, we created 398 SSR markers from BAC end and EST sequences. An analysis of 187 diploid lines from 56 locations with 43 SSR markers showed considerable genotypic diversity. There was some correlation between SSR genotypes and broad geographic regions, but there was also a high level of genotypic diversity at individual locations. Phenotypic analysis of this new germplasm resource revealed considerable variation in flowering time, seed size, and plant architecture. The inbreeding nature of Brachypodium was confirmed by an extremely high level of homozygosity in wild plants and a lack of cross-pollination under laboratory conditions. Conclusion: Taken together, the inbreeding nature and genotypic diversity observed at individual locations suggest a significant amount of long-distance seed dispersal. The resources developed in this study are freely available to the research community and will facilitate experimental applications based on natural diversity

Selection and phenotypic characterization of a core collection of <i>Brachypodium distachyon</i> inbred lines

BACKGROUND: The model grass Brachypodium distachyon is increasingly used to study various aspects of grass biology. A large and genotypically diverse collection of B. distachyon germplasm has been assembled by the research community. The natural variation in this collection can serve as a powerful experimental tool for many areas of inquiry, including investigating biomass traits. RESULTS: We surveyed the phenotypic diversity in a large collection of inbred lines and then selected a core collection of lines for more detailed analysis with an emphasis on traits relevant to the use of grasses as biofuel and grain crops. Phenotypic characters examined included plant height, growth habit, stem density, flowering time, and seed weight. We also surveyed differences in cell wall composition using near infrared spectroscopy (NIR) and comprehensive microarray polymer profiling (CoMPP). In all cases, we observed extensive natural variation including a two-fold variation in stem density, four-fold variation in ferulic acid bound to hemicellulose, and 1.7-fold variation in seed mass. CONCLUSION: These characterizations can provide the criteria for selecting diverse lines for future investigations of the genetic basis of the observed phenotypic variation

A Program for Handling Map Projections of Small Scale Geospatial Raster Data

Scientists routinely accomplish small-scale geospatial modeling using raster datasets of global extent. Such use often requires the projection of global raster datasets onto a map or the reprojection from a given map projection associated with a dataset. The distortion characteristics of these projection transformations can have significant effects on modeling results. Distortions associated with the reprojection of global data are generally greater than distortions associated with reprojections of larger-scale, localized areas. The accuracy of areas in projected raster datasets of global extent is dependent on resolution. To address these problems of projection and the associated resampling that accompanies it, methods for framing the transformation space, direct point-to-point transformations rather than gridded transformation spaces, a solution to the wrap-around problem, and an approach to alternative resampling methods are presented. The implementations of these methods are provided in an open source software package called MapImage (or mapIMG, for short), which is designed to function on a variety of computer architectures

Unrooted neighbor joining consensus tree of 19 lines based on 1,000 shared allele bootstrap trees constructed using 10 SSR markers.

<p>Bootstrap values greater than 400 are shown.</p

Growth habit and chromosomes.

<p>(a) Flowering plants in the growth chamber. The accessions are, from left to right, PI296842, PI297868 and PI204863. Note the variation in height. The pots are 10 cm tall. (b) Close up of flowers. Note the fully exerted anthers and stigmas. (c) Bench full of transgenic plants surrounding a non-transgenic plant to determine pollen flow. The red arrow is pointing to a red stake in the pot of the non-transgenic plant. (d-f) Mitotic chromosome spreads of the indicated accessions. Scale bars = 5 µm.</p

<i>Brachypodium</i><i> sylvaticum</i>, a Model for Perennial Grasses: Transformation and Inbred Line Development

<div><p>Perennial species offer significant advantages as crops including reduced soil erosion, lower energy inputs after the first year, deeper root systems that access more soil moisture, and decreased fertilizer inputs due to the remobilization of nutrients at the end of the growing season. These advantages are particularly relevant for emerging biomass crops and it is projected that perennial grasses will be among the most important dedicated biomass crops. The advantages offered by perennial crops could also prove favorable for incorporation into annual grain crops like wheat, rice, sorghum and barley, especially under the dryer and more variable climate conditions projected for many grain-producing regions. Thus, it would be useful to have a perennial model system to test biotechnological approaches to crop improvement and for fundamental research. The perennial grass <i>Brachypodium</i><i>sylvaticum</i> is a candidate for such a model because it is diploid, has a small genome, is self-fertile, has a modest stature, and short generation time. Its close relationship to the annual model <i>Brachypodium</i><i>distachyon</i> will facilitate comparative studies and allow researchers to leverage the resources developed for <i>B</i><i>. distachyon</i>. Here we report on the development of two keystone resources that are essential for a model plant: high-efficiency transformation and inbred lines. Using <i>Agrobacterium tumefaciens</i>-mediated transformation we achieved an average transformation efficiency of 67%. We also surveyed the genetic diversity of 19 accessions from the National Plant Germplasm System using SSR markers and created 15 inbred lines.</p> </div

Approximate collection locations for the lines used in this study.

<p>All locations were inferred from the collection information found in the Plant Inventory accessed through the GRIN database. For most lines the town nearest the collection site was mapped. Locations 20-22 were mapped using the GPS coordinates of the collection locations. For location 4 and 12 only the country of origin was listed so the map location indicated is the approximate center of the country. See Table 1 for location number to accession assignments. For complete collection data see Figure S1.</p

Tissue culture stages of inbred line Ain-1.

<p>(a) Embryos dissected from immature seeds. The five embryos on the left will produce high quality embryogenic callus. The embryo on the right is slightly too large. Note that it is also more opaque. The scale bar is 1 mm. (b) Appearance of high-quality embryogenic callus. Note the structures that look like embryos. Scale bar is 5 mm. (c) Callus on dry filter paper after removal from <i>Agrobacterium</i> suspension. The plate is 10 cm in diameter. (d) Transgenic plantlets regenerating from callus. Note the presence of darker non-transgenic callus that is dying. Scale bar is 1cm. (e) GUS positive (top) and negative (bottom) plants representative of the transgenic outcrossed progeny and non-transgenic selfed progeny of the non-transgenic plants used to determine outcrossing frequency.</p