B. Shaun Bushman

Orchardgrass (Dactylis glomerata L.) EST and SSR marker development, annotation, and transferability.

Orchardgrass, or cocksfoot [Dactylis glomerata (L.)], has been naturalized on nearly every continent and is a commonly used species for forage and hay production. All major cultivated varieties of orchardgrass are autotetraploid, and few tools or information are available for functional and comparative genetic analyses and improvement of the species. To improve the genetic resources for orchardgrass, we have developed an EST library and SSR markers from salt, drought, and cold stressed tissues. The ESTs were bi-directionally sequenced from clones and combined into 17,373 unigenes. Unigenes were annotated based on putative orthology to genes from rice, Triticeae grasses, other Poaceae, Arabidopsis, and the non-redundant database of the NCBI. Of 1,162 SSR markers developed, approximately 80% showed amplification products across a set of orchardgrass germplasm, and 40% across related Festuca and Lolium species. When orchardgrass subspecies were genotyped using 33 SSR markers their within-accession similarity values ranged from 0.44 to 0.71, with Mediterranean accessions having a higher similarity. The total number of genotyped bands was greater for tetraploid accessions compared to diploid accessions. Clustering analysis indicated grouping of Mediterranean subspecies and central Asian subspecies, while the D. glomerata ssp. aschersoniana was closest related to three cultivated varieties.

Development and annotation of perennial Triticeae ESTs and SSR markers.

Triticeae contains hundreds of species of both annual and perennial types. Although substantial genomic tools are available for annual Triticeae cereals such as wheat and barley, the perennial Triticeae lack sufficient genomic resources for genetic mapping or diversity research. To increase the amount of sequence information available in the perennial Triticeae, three expressed sequence tag (EST) libraries were developed and annotated for Pseudoroegneria spicata, a mixture of both Elymus wawawaiensis and E. lanceolatus, and a Leymus cinereus x L. triticoides interspecific hybrid. The ESTs were combined into unigene sets of 8 780 unigenes for P. spicata, 11 281 unigenes for Leymus, and 7 212 unigenes for Elymus. Unigenes were annotated based on putative orthology to genes from rice, wheat, barley, other Poaceae, Arabidopsis, and the non-redundant database of the NCBI. Simple sequence repeat (SSR) markers were developed, tested for amplification and polymorphism, and aligned to the rice genome. Leymus EST markers homologous to rice chromosome 2 genes were syntenous on Leymus homeologous groups 6a and 6b (previously 1b), demonstrating promise for in silico comparative mapping. All ESTs and SSR markers are available on an EST information management and annotation database (http://titan.biotec.uiuc.edu/triticeae/).

Genes controlling plant growth habit in Leymus (Triticeae): maize barren stalk1 (ba1), rice lax panicle, and wheat tiller inhibition (tin3) genes as possible candidates

Leymus cinereus and L. triticoides are large caespitose and rhizomatous perennial grasses, respectively. Previous studies detected quantitative trait loci (QTL) controlling rhizome spreading near the viviparous1 (vp1) gene markers on linkage groups LG3a and LG3b in two families, TTC1 and TTC2, derived from Leymus triticoides × Leymus cinereus hybrids. The wheat tiller inhibition gene (tin3) is located on Triticum monococcum chromosome 3 AmL near vp1. Triticeae group 3 is reportedly collinear with rice chromosome 1, which also contains the maize barren stalk1 and rice lax branching orthogene near vp1. However, previous studies lacked cross-species markers for comparative mapping and showed possible rearrangements of Leymus group 3 in wheat-Leymus racemosus chromosome addition lines. Here, we developed expressed sequence tag (EST) markers from Leymus tiller and rhizomes and mapped sequences aligned to rice chromosome 1. Thirty-eight of 44 informative markers detected loci on Leymus LG3a and LG3b that were collinear with homoeologous sequences on rice chromosome 1 and syntenous in homoeologous group 3 wheat-Leymus and wheat-Thinopyrum addition lines. A SCARECROW-like GRAS-family transcription factor candidate gene was identified in the Leymus EST library, which aligns to the Leymus chromosome group 3 growth habit QTL and a 324-kb rice chromosome 1 region thought to contain the wheat tin3 gene.

Phenotypic and Genetic Characterization of Western Prairie Clover Collections From the Western United States

Abstract Few North American legumes are available for rangeland revegetation in the semiarid western United States. Western prairie clover (Dalea ornata [Douglas ex Hook.] Eaton & J. Wright) is a perennial legume with desirable forage characteristics and is distributed in the northern Great Basin, Snake River Basin, and southern Columbia Plateau. Understanding the genetic and ecotypic variability of this species is a prerequisite for developing populations suitable for revegetation purposes. To address this need, we established two common-garden plots of western prairie clover from 22 sites in Idaho, Oregon, and Washington. Significant variation was detected among the collections for all traits measured. Among the measured traits, flowering date was correlated with collection-site temperature and elevation. Population structure estimates from 474 amplified-fragment length polymorphism markers resulted in two distinct, genetically differentiated groups and a third admixed group, and flowering date played a significant role in discriminating those genetic-based groupings of collections. Positive correlations were observed between phenotypic and genetic distance matrices (r  =  0.33, P  =  0.005), phenotypic and geographic distance matrices (r  =  0.35, P  =  0.002), and genetic and geographic distance matrices (r  =  0.31, P  =  0.009). Based on these results, we recommend that two germplasm sources of western prairie clover be developed for use across the collection area, one from the Deschutes River region and the other encompassing Idaho, Washington, and eastern Oregon collection sites.

Notice of Release of NBR-1 Germplasm Basalt Milkvetch

A selected-class pre-variety germplasm of basalt milkvetch (Astragalus filipes Torr. ex A. Gray [Fabaceae]) has been released for reclamation, rehabilitation, and restoration of semiarid rangelands in the northern Great Basin Region of the western US.Johnson DA, Jones TA, Connors KJ, Bhattarai K, Bushman BS, Jensen KB. 2008. Notice of release of NBR-1 Germplasm basalt milkvetch. Native Plants Journal 9(2):127–132.

Genome evolution of intermediate wheatgrass as revealed by EST-SSR markers developed from its three progenitor diploid species

Intermediate wheatgrass (Thinopyrum intermedium (Host) Barkworth & D.R. Dewey), a segmental autoallohexaploid (2n = 6x = 42), is not only an important forage crop but also a valuable gene reservoir for wheat (Triticum aestivum L.) improvement. Throughout the scientific literature, there continues to be disagreement as to the origin of the different genomes in intermediate wheatgrass. Genotypic data obtained from newly developed EST-SSR primers derived from the putative progenitor diploid species Pseudoroegneria spicata (Pursh) Á. Löve (St genome), Thinopyrum bessarabicum (Savul. & Rayss) Á. Löve (J = J(b) = E(b)), and Thinopyrum elongatum (Host) D. Dewey (E = J(e) = E(e)) indicate that the V genome of Dasypyrum (Coss. & Durieu) T. Durand is not one of the three genomes in intermediate wheatgrass. Based on all available information in the literature and findings in this study, the genomic designation of intermediate wheatgrass should be changed to J(vs)J(r)St, where J(vs) and J(r) represent ancestral genomes of present-day J(b) of Th. bessarabicum and J(e) of Th. elongatum, with J(vs) being more ancient. Furthermore, the information suggests that the St genome in intermediate wheatgrass is most similar to the present-day St found in diploid species of Pseudoroegneria from Eurasia.

Seed Dormancy Mechanisms in Basalt Milkvetch and Western Prairie Clover

ABSTRACT A greater diversity of native legumes and forbs is desirable for rangeland restoration practice in the Intermountain Region of the western United States. But for such diversity to materialize in the seed marketplace and to be effective in restoration practice, seeds that germinate reliably in seed fields and on restoration sites are needed. We measured germination response of two native legumes, basalt milkvetch (Astragalus filipes Torr. ex A. Gray) and western prairie clover (Dalea ornata [Douglas] Eaton & Wright), after eight germination treatments. Treatments were a factorial combination of 1) seed scarification with sandpaper (or unscarified), 2) a substrate of moist sand (or blotter paper), and 3) a 3-wk prechill at 5° (or nonprechilled). Cumulative germination increased linearly throughout the 10-wk course of the experiment for all treatment combinations in both species. Scarification increased germination of western prairie clover, but prechilling and substrate had no effect. In contrast, prechilling, scarification, and a sand substrate all increased germination of basalt milkvetch. Hence, for this species the prechilled/scarified/sand treatment combination displayed the numerically highest germination for all 10 wk (30–43%), and the nonprechilled/unscarified/blotter paper treatment combination always germinated lowest (1–3%). Results were consistent with physical dormancy (hard-seededness) limiting germination of western prairie clover and combinational dormancy (i.e., co-occurrence of physical and physiological dormancy) limiting germination of basalt milkvetch. Of the two species, we have found basalt milkvetch to be the more difficult to establish from seed. By prechilling acid-scarified seed in moist sand, basalt milkvetch was successfully established in two field trials seeded in mid-April. Nonprechilled mechanically (sandpaper) scarified seed germinated as high as prechilled acid-scarified seed. By scarifying and prechilling basalt milkvetch seed to address physical and physiological dormancy mechanisms, respectively, this seed-treatment protocol may be “scaled up” to produce large quantities of germinable seed.

Relationships among seed quality characteristics in a collection of western wheatgrass germplasms

Although western wheatgrass [Pascopyrum smithii (Rydb.)] is an important perennial grass species for agriculture and conservation management in central and western North America, its lack of adequate seed production and seedling vigor limits its effectiveness. To address the weaknesses a study was conducted to assess rhizome spread, seed production, seed weight, germination percentage, and emergence rate of seed produced from 48 western wheatgrass cultivars and germplasm accessions at a field site near Nephi, UT, USA during 2007 and 2008. The western wheatgrass cultivars had approximately two times higher seed production than the germplasm accessions during both 2007 and 2008 and also had higher seed weight in 2007 and emergence rate in 2008. The germplasm accessions had higher seed weight in 2008. For the remaining traits there were no differences among the different germplasm sources. Based on principle component analysis a subset of cultivars and germplasm accessions with high seed production and emergence rate were identified that could be used to produce improved cultivars and germplasms. There was little evidence of strong relationship between geographic, genetic, and phenotypic distances among the various lines examined. Additionally, based on genetic marker data, a subset of lines was grouped into three populations. Based on these results, selection among lines could occur to maximize agricultural performance regardless of site of origin, or within population selection could be practiced to meet conservation goals of minimizing hybridization among populations.

Utah Lotus: North American Legume for Rangeland Revegetation in the Southern Great Basin and Colorado Plateau☆,☆☆

ABSTRACT Utah lotus (Lotus utahensis Ottley) is a North American leguminous forb that may hold promise for rangeland revegetation in the western United States for diversifying planting mixtures, attracting pollinators, providing high-quality forage, and expanding habitats for insects needed by sage-grouse chicks. Fourteen wildland seed collections of Utah lotus originating from Nevada and Utah were assessed for genetic variation of a wide range of phenotypic traits and genetic relationships. Population structure estimates defined by 552 amplified fragment length polymorphism (AFLP) markers identified three primary subgroups within the Utah lotus collections, which corresponded to their geographic origin. Two collections of Utah lotus (LU-5 and LU-20) were among the top-performing collections for the phenotypic traits examined, including dry-matter yield, pod production, number of stems, canopy height, and persistence. No significant Pearsons correlations or canonical correlations were observed among the phenotypic traits and environmental characteristics at the collection sites. Significant correlations were detected between genetic and geographic matrices, and phenotypic and geographic distance matrices (r = 0.89, P = 0.001 and r = 0.24, P = 0.04, respectively). Condensed tannin (CT) contents of Utah lotus were between 146 and 199 g kg-1 dry matter, which was nearly 10 times higher than CT content of birdsfoot trefoil (Lotus corniculatus L.) with 17.3 g kg-1 dry matter. Because of our phenotypic and genotypic evaluations, one pooled germplasm source of Utah lotus comprising collections LU-5 and LU-20 could be developed for use in rangeland revegetation in the southern Great Basin and Colorado Plateau.

Effects of Selection for Seedling Vigor on the Genetic Variation in Leymus cinereus

ABSTRACT Basin wildrye (Leymus cinereus [Scribn. & Merr. Á. Löve]) is a perennial grass native to western regions of North America. Despite its importance for rangelands, stand establishment of basin wildrye is difficult due to its poor seedling vigor. We undertook to increase the seedling vigor of the basinwildrye cultivar “Trailhead” by using selection for emergence fromdeep seeding depth. We carried out two cycles of selection in two select populations and included two random populations, in which no direct selection occurred. We characterized the indirection effect of the selection on biomass, seed production, and stand percentage in these populations under field conditions. We used amplified fragment length polymorphic (AFLP)markers to identify regions of the genome associated with the selection by identifying allele frequency changes between the base population and the select and random populations. The second cycle select population and the first cycle random population possessed the highest total emergence from deep seeding (60% and 59%, respectively) compared with the base population (26%). The field evaluations showed no differences in genetic variation among the base, select, and random populations for biomass, seed production, and stand percentage. On the basis of the analysis of the AFLPmarkers, diversity increased slightly among the random populations and decreased slightly among the select populations. In the select populations, band frequencies increased for aggcac403, actcag185, and aggcac208. The band frequencies of aggctg212 and actctc66 decreased in both random and selected cycles. The results indicate that targeted selection for trait improvement in this native grass can be successfully completed with minimal effect on population genetic diversity.