QTL for seed shattering and threshability in intermediate wheatgrass align closely with well-studied orthologs from wheat, barley, and rice.

Perennial grain crops have the potential to improve agricultural sustainability but few existing species produce sufficient grain yield to be economically viable. The outcrossing, allohexaploid, and perennial forage species intermediate wheatgrass (IWG) [Thinopyrum intermedium (Host) Barkworth & D. R. Dewey] has shown promise in undergoing direct domestication as a perennial grain crop using phenotypic and genomic selection. However, decades of selection will be required to achieve yields on par with annual small-grain crops. Marker-aided selection could accelerate progress if important genomic regions associated with domestication were identified. Here we use the IWG nested association mapping (NAM) population, with 1,168 F1 progeny across 10 families to dissect the genetic control of brittle rachis, floret shattering, and threshability. We used a genome-wide association study (GWAS) with 8,003 single nucleotide polymorphism (SNP) markers and linkage mapping-both within-family and combined across families-with a robust phenotypic dataset collected from four unique year-by-location combinations. A total of 29 quantitative trait loci (QTL) using GWAS and 20 using the combined linkage analysis were detected, and most large-effect QTL were in common across the two analysis methods. We reveal that the genetic control of these traits in IWG is complex, with significant QTL across multiple chromosomes, sometimes within and across homoeologous groups and effects that vary depending on the family. In some cases, these QTL align within 216 bp to 31 Mbp of BLAST hits for known domestication genes in related species and may serve as precise targets of selection and directions for further study to advance the domestication of IWG.

DNA sequence-based mapping and comparative genomics of the St genome of Pseudoroegneria spicata (Pursh) Á. Löve versus wheat (Triticum aestivum L.) and barley (Hordeum vulgare L.).

Bluebunch wheatgrass (referred to as BBWG) [Pseudoroegneria spicata (Pursh) Á. Löve] is an important rangeland Triticeae grass used for forage, conservation, and restoration. This diploid has the basic St genome that occurs also in many polyploid Triticeae species, which serve as a gene reservoir for wheat improvement. Until now, the St genome in diploid species of Pseudoroegneria has not been mapped. Using a double-cross mapping populations, we mapped 230 expressed sequence tag derived simple sequence repeat (EST-SSR) and 3468 genotyping-by-sequencing (GBS) markers to 14 linkage groups (LGs), two each for the seven homologous groups of the St genome. The 227 GBS markers of BBWG that matched those in a previous study helped identify the unclassified seven LGs of the St sub-genome among 21 LGs of Thinopyrum intermedium (Host) Barkworth & D.R. Dewey. Comparisons of GBS sequences in BBWG to whole-genome sequences in bread wheat (Triticum aestivum L.) and barley (Hordeum vulgare L.) revealed that the St genome shared a homology of 35% and 24%, a synteny of 86% and 84%, and a collinearity of 0.85 and 0.86, with ABD and H, respectively. This first-draft molecular map of the St genome will be useful in breeding cereal and forage crops.

Differential transferability of EST-SSR primers developed from the diploid species Pseudoroegneria spicata, Thinopyrum bessarabicum, and Thinopyrum elongatum.

Simple sequence repeat technology based on expressed sequence tag (EST-SSR) is a useful genomic tool for genome mapping, characterizing plant species relationships, elucidating genome evolution, and tracing genes on alien chromosome segments. EST-SSR primers developed from three perennial diploid species of Triticeae, Pseudoroegneria spicata (Pursh) Á. Löve (having St genome), Thinopyrum bessarabicum (Savul. & Rayss) Á. Löve (Jb = Eb = J), and Thinopyrum elongatum (Host) D.R. Dewey (Je = Ee = E), were used to produce amplicons in these three species to (i) assess relative transferability, (ii) identify polymorphic species-specific markers, and (iii) determine genome relationships among the three species. Because of the close relationship between Jb and Je genomes, EST-SSR primers derived from Th. bessarabicum and Th. elongatum had greater transferability to each other than those derived from the St-genome P. spicata. A large number of polymorphic species- and genome-specific EST-SSR amplicons were identified that will be used for construction of genetic maps of these diploid species, and tracing economically useful genes in breeding or gene transfer programs in various species of Triticeae.

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.

Genetic control of rhizomes and genomic localization of a major-effect growth habit QTL in perennial wildrye.

Rhizomes are prostrate subterranean stems that provide primitive mechanisms of vegetative dispersal, survival, and regrowth of perennial grasses and other monocots. The extent of rhizome proliferation varies greatly among grasses, being absent in cereals and other annuals, strictly confined in caespitose perennials, or highly invasive in some perennial weeds. However, genetic studies of rhizome proliferation are limited and genes controlling rhizomatous growth habit have not been elucidated. Quantitative trait loci (QTLs) controlling rhizome spreading were compared in reciprocal backcross populations derived from hybrids of rhizomatous creeping wildrye (Leymus triticoides) and caespitose basin wildrye (L. cinereus), which are perennial relatives of wheat. Two recessive QTLs were unique to the creeping wildrye backcross, one dominant QTL was unique to the basin wildrye backcross, and one additive QTL was detectable in reciprocal backcrosses with high log odds (LOD = 31.6) in the basin wildrye background. The dominant QTL located on linkage group (LG)-2a was aligned to a dominant rhizome orthogene (Rhz3) of perennial rice (Oryza longistamina) and perennial sorghum (Sorghum propinquum). Nonparametric 99 % confidence bounds of the 31.6-LOD QTL were localized to a distal 3.8-centiMorgan region of LG-6a, which corresponds to a 0.7-Mb region of Brachypodium Chromosome 3 containing 106 genes. An Aux/IAA auxin signal factor gene was located at the 31.6-LOD peak, which could explain the gravitropic and aphototropic behavior of rhizomes. Findings elucidate genetic mechanisms controlling rhizome development and architectural growth habit differences among plant species. Results have possible applications to improve perennial forage and turf grasses, extend the vegetative life cycle of annual cereals, such as wheat, or control the invasiveness of highly rhizomatous weeds such as quackgrass (Elymus repens).

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/).