Molecular discrimination of tall fescue morphotypes in association with Festuca relatives.

Tall fescue (Festuca arundinacea Schreb.) is an important cool-season perennial grass species used as forage and turf, and in conservation plantings. There are three morphotypes in hexaploid tall fescue: Continental, Mediterranean and Rhizomatous. This study was conducted to develop morphotype-specific molecular markers to distinguish Continental and Mediterranean tall fescues, and establish their relationships with other species of the Festuca genus for genomic inference. Chloroplast sequence variation and simple sequence repeat (SSR) polymorphism were explored in 12 genotypes of three tall fescue morphotypes and four Festuca species. Hypervariable chloroplast regions were retrieved by using 33 specifically designed primers followed by sequencing the PCR products. SSR polymorphism was studied using 144 tall fescue SSR primers. Four chloroplast (NFTCHL17, NFTCHL43, NFTCHL45 and NFTCHL48) and three SSR (nffa090, nffa204 and nffa338) markers were identified which can distinctly differentiate Continental and Mediterranean morphotypes. A primer pair, NFTCHL45, amplified a 47 bp deletion between the two morphotypes is being routinely used in the Noble Research Institute's core facility for morphotype discrimination. Both chloroplast sequence variation and SSR diversity showed a close association between Rhizomatous and Continental morphotypes, while the Mediterranean morphotype was in a distant clade. F. pratensis and F. arundinacea var. glaucescens, the P and G1G2 genome donors, respectively, were grouped with the Continental clade, and F. mairei (M1M2 genome) grouped with the Mediterranean clade in chloroplast sequence variation, while both F. pratensis and F. mairei formed independent clade in SSR analysis. Age estimation based on chloroplast sequence variation indicated that the Continental and Mediterranean clades might have been colonized independently during 0.65 ± 0.06 and 0.96 ± 0.1 million years ago (Mya) respectively. The findings of the study will enhance tall fescue breeding for persistence and productivity.

Evolution of the Grain Dispersal System in Barley.

About 12,000 years ago in the Near East, humans began the transition from hunter-gathering to agriculture-based societies. Barley was a founder crop in this process, and the most important steps in its domestication were mutations in two adjacent, dominant, and complementary genes, through which grains were retained on the inflorescence at maturity, enabling effective harvesting. Independent recessive mutations in each of these genes caused cell wall thickening in a highly specific grain "disarticulation zone," converting the brittle floral axis (the rachis) of the wild-type into a tough, non-brittle form that promoted grain retention. By tracing the evolutionary history of allelic variation in both genes, we conclude that spatially and temporally independent selections of germplasm with a non-brittle rachis were made during the domestication of barley by farmers in the southern and northern regions of the Levant, actions that made a major contribution to the emergence of early agrarian societies.

Linkage Maps of a Mediterranean × Continental Tall Fescue Population and their Comparative Analysis with Other Poaceae Species.

Temperate grasses belonging to the Festuca-Lolium complex are important throughout the world in pasture and grassland agriculture. Tall fescue (Festuca arundinacea Schreb.) is the predominant species in the United States, covering approximately 15 million ha. Tall fescue has distinctive morphotypes, two of which are Continental (summer active) and Mediterranean (summer semidormant). This is the first report of a linkage map created for Mediterranean tall fescue, while updating the Continental map with additional simple sequence repeat and sequence-tagged site markers. Additionally, this is the first time that diversity arrays technology (DArT) markers were used in the construction of a tall fescue map. The male parent (Continental), R43-64, map consisted of 594 markers arranged in 22 linkage groups (LGs) and covered a total of 1577 cM. The female parent (Mediterranean), 103-2, map was shorter (1258 cM) and consisted of only 208 markers arranged in 29 LGs. Marker densities for R43-64 and 103-2 were 2.65 and 6.08 cM per marker, respectively. When compared with the other Poaceae species, meadow fescue (F. pratensis Huds.), annual ryegrass (L. multiflorum Lam.), perennial ryegrass (L. perenne L.), Brachypodium distachyon (L.) Beauv., and barley (Hordeum vulgare L.), a total of 171 and 98 orthologous or homologous sequences, identified by DArT analysis, were identified in R43-64 and 103-2, respectively. By using genomic in situ hybridization, we aimed to identify potential progenitors of both morphotypes. However, no clear conclusion on genomic constitution was reached. These maps will aid in the search for quantitative trait loci of various traits as well as help define and distinguish genetic differences between the two morphotypes.

Fine genetic mapping of greenbug aphid-resistance gene Gb3 in Aegilops tauschii.

The greenbug, Schizaphis graminum (Rondani), is an important aphid pest of small grain crops especially wheat (Triticum aestivum L., 2n = 6x = 42, genomes AABBDD) in many parts of the world. The greenbug-resistance gene Gb3 originated from Aegilops tauschii Coss. (2n = 2x = 14, genome D(t)D(t)) has shown consistent and durable resistance against prevailing greenbug biotypes in wheat fields. We previously mapped Gb3 in a recombination-rich, telomeric bin of wheat chromosome arm 7DL. In this study, high-resolution genetic mapping was carried out using an F(2:3) segregating population derived from two Ae. tauschii accessions, the resistant PI 268210 (original donor of Gb3 in the hexaploid wheat germplasm line 'Largo') and susceptible AL8/78. Molecular markers were developed by exploring bin-mapped wheat RFLPs, SSRs, ESTs and the Ae. tauschii physical map (BAC contigs). Wheat EST and Ae. tauschii BAC end sequences located in the deletion bin 7DL3-0.82-1.00 were used to design STS (sequence tagged site) or CAPS (Cleaved Amplified Polymorphic Sequence) markers. Forty-five PCR-based markers were developed and mapped to the chromosomal region spanning the Gb3 locus. The greenbug-resistance gene Gb3 now was delimited in an interval of 1.1 cM by two molecular markers (HI067J6-R and HI009B3-R). This localized high-resolution genetic map with markers closely linked to Gb3 lays a solid foundation for map based cloning of Gb3 and marker-assisted selection of this gene in wheat breeding.

BAC libraries from wheat chromosome 7D: efficient tool for positional cloning of aphid resistance genes.

Positional cloning in bread wheat is a tedious task due to its huge genome size and hexaploid character. BAC libraries represent an essential tool for positional cloning. However, wheat BAC libraries comprise more than million clones, which makes their screening very laborious. Here, we present a targeted approach based on chromosome-specific BAC libraries. Such libraries were constructed from flow-sorted arms of wheat chromosome 7D. A library from the short arm (7DS) consisting of 49,152 clones with 113 kb insert size represented 12.1 arm equivalents whereas a library from the long arm (7DL) comprised 50,304 clones of 116 kb providing 14.9x arm coverage. The 7DS library was PCR screened with markers linked to Russian wheat aphid resistance gene DnCI2401, the 7DL library was screened by hybridization with a probe linked to greenbug resistance gene Gb3. The small number of clones combined with high coverage made the screening highly efficient and cost effective.

A phylogenetic analysis based on nucleotide sequence of a marker linked to the brittle rachis locus indicates a diphyletic origin of barley.

BACKGROUND AND AIMS: Barley (Hordeum vulgare ssp. vulgare) cultivation started between 9500 and 8400 years ago, and was a major part of ancient agriculture in the Near East. The brittle rachis is a critical trait in the domestication process. METHODS: A DNA sequence closely linked to the brittle rachis complex was amplified and resequenced in a collection of cultivated barleys, wild barleys (H. vulgare ssp. spontaneum) and weedy brittle rachis varieties (H. vulgare ssp. vulgare var. agriocrithon). The sequence was used to construct a phylogenetic tree. KEY RESULTS: The phylogeny separated the W- (btr1-carrying) from the E- (btr2-carrying) cultivars. The wild barleys had a high sequence diversity and were distributed throughout the W- and E-clades. Some of the Tibetan var. agriocrithon lines were closely related to the E-type and others to the W-type cultivated barleys, but an Israeli var. agriocrithon line has a complex origin. CONCLUSIONS: The results are consistent with a diphyletic origin of barley. The W- and E-type cultivars are assumed to have evolved from previously diverged wild barley via independent mutations at Btr1 and Btr2.

Six-rowed barley originated from a mutation in a homeodomain-leucine zipper I-class homeobox gene.

Increased seed production has been a common goal during the domestication of cereal crops, and early cultivators of barley (Hordeum vulgare ssp. vulgare) selected a phenotype with a six-rowed spike that stably produced three times the usual grain number. This improved yield established barley as a founder crop for the Near Eastern Neolithic civilization. The barley spike has one central and two lateral spikelets at each rachis node. The wild-type progenitor (H. vulgare ssp. spontaneum) has a two-rowed phenotype, with additional, strictly rudimentary, lateral rows; this natural adaptation is advantageous for seed dispersal after shattering. Until recently, the origin of the six-rowed phenotype remained unknown. In the present study, we isolated vrs1 (six-rowed spike 1), the gene responsible for the six-rowed spike in barley, by means of positional cloning. The wild-type Vrs1 allele (for two-rowed barley) encodes a transcription factor that includes a homeodomain with a closely linked leucine zipper motif. Expression of Vrs1 was strictly localized in the lateral-spikelet primordia of immature spikes, suggesting that the VRS1 protein suppresses development of the lateral rows. Loss of function of Vrs1 resulted in complete conversion of the rudimentary lateral spikelets in two-rowed barley into fully developed fertile spikelets in the six-rowed phenotype. Phylogenetic analysis demonstrated that the six-rowed phenotype originated repeatedly, at different times and in different regions, through independent mutations of Vrs1.