Free Threshing Research Articles

Genetic constitution and variability in synthetic populations of intermediate wheatgrass, an outcrossing perennial grain crop.

Intermediate wheatgrass (IWG) is a perennial grass that produces nutritious grain while offering substantial ecosystem services. Commercial varieties of this crop are mostly synthetic panmictic populations that are developed by intermating a few selected individuals. As development and generation advancement of these synthetic populations is a multiyear process, earlier synthetic generations are tested by the breeders and subsequent generations are released to the growers. A comparison of generations within IWG synthetic cultivars is currently lacking. In this study, we used simulation models and genomic prediction to analyze population differences and trends of genetic variance in 4 synthetic generations of MN-Clearwater, a commercial cultivar released by the University of Minnesota. Little to no differences were observed among the 4 generations for population genetic, genetic kinship, and genome-wide marker relationships measured via linkage disequilibrium. A reduction in genetic variance was observed when 7 parents were used to generate synthetic populations while using 20 led to the best possible outcome in determining population variance. Genomic prediction of plant height, free threshing ability, seed mass, and grain yield among the 4 synthetic generations showed a few significant differences among the generations, yet the differences in values were negligible. Based on these observations, we make 2 major conclusions: (1) the earlier and latter synthetic generations of IWG are mostly similar to each other with minimal differences and (2) using 20 genotypes to create synthetic populations is recommended to sustain ample genetic variance and trait expression among all synthetic generations.

Agropastoral and dietary practices of the northern Levant facing Late Holocene climate and environmental change: Isotopic analysis of plants, animals and humans from Bronze to Iron Age Tell Tweini.

One of the largest isotopic datasets of the ancient Eastern Mediterranean region is evaluated, based on plants (n = 410), animals (n = 210) and humans (n = 16) from Tell Tweini (Syria). Diachronic analysis of plant and faunal specimens from four main periods of occupation: Early Bronze Age (2600-2000 BC), Middle Bronze Age (2000-1600 BC), Late Bronze Age (1600-1200 BC) and Iron Age (1200-333 BC) were investigated. Mean Δ13C results from seven plant species reveal emmer and free threshing wheat, olives, bitter vetch, rye grass and barley were adequately or well-watered during all periods of occupation. The grape Δ13C results suggest excellent growing conditions and particular care for its cultivation. The δ15N results indicate that especially the emmer and free threshing wheats received some manure inputs throughout the occupation sequence, while these were likely further increased during the Iron Age, encompassing also the olive groves and grape vineyards. Generally, domestic animals (cattle, sheep, goats) had C3 terrestrial diets and were kept together in similar environments. However, some animals consumed significant amounts of marine or C4 plants, possibly from disturbed habitats due to land use pressure or salt tolerant grasses and shrubs from wetland environments, which were recorded in the direct vicinity of the site. Middle Bronze Age humans consumed a C3 terrestrial diet with no measurable input from C4, freshwater or marine protein sources. Interestingly, the human diet was relatively low in animal protein and appears comparable to what is considered today a typical Mediterranean diet consisting of bread (wheat/barley), olives, grapes, pulses, dairy products and small amounts of meat. The combined isotopic analysis of plants, animals and humans from Tell Tweini represents unbroken links in the food chain which create unparalleled opportunities to enhance our current understanding of environmental conditions, climate change and lifeways in past populations from the Eastern Mediterranean.

Genomic prediction enables rapid selection of high-performing genets in an intermediate wheatgrass breeding program.

In an era of constrained and depleted natural resources, perennial grains could provide sustainable food production along with beneficial ecosystem services like reduced erosion and increased atmospheric carbon capture. Intermediate wheatgrass (IWG) [Thinopyrum intermedium (Host) Barkworth & D. R. Dewey subsp. intermedium] has been undergoing continuous breeding for domestication to develop a perennial grain crop since the 1980s. As a perennial, IWG has required 2-5 yr per selection generation, but starting in 2017, genomic selection (GS) was initiated in the breeding program at The Land Institute, Salina, KS (TLI), enabling one complete cycle per year. For each cycle, ∼4,000 seedlings were profiled using genotyping-by-sequencing (GBS) and genomic estimated breeding values (GEBVs) were calculated. Selection based on GEBVs identified ∼100 individuals to advance as parents each generation, while validation populations of 1,000-1,200 genets for GS model training were also selected using the genomic relationship matrix to represent genetic diversity in each cycle. The selected parents were randomly intermated in a greenhouse crossing block to form the subsequent cycle, while the validation populations were transplanted to irrigated and nonirrigated field sites for phenotypic evaluations in the following years. For priority breeding traits of seed mass, free threshing, and nonshattering, correlations between predicted values and observed data were >.5. The realized selection differential ranged from 11-23% for selected traits, and the expected genetic gains for these traits, including spike yield, ranged from 6 to 14% per year. Genomic selection is a powerful tool to speed the domestication and development of IWG and other perennial crops with extended breeding timelines.

A novel wheat q’ allele identified by forward genetic in silico TILLING

The major wheat domestication allele Q (encoding an APETALA2 like transcription factor) is responsible for the free threshing and square-headed spikes of modern wheat. Wild type q and null q’ alleles cause a reversal to the speltoid phenotype. Q pleiotropically affects additional yield and quality traits so genetic variation in Q and its interaction partners remain important for crop improvement. Here we report the discovery of a novel q’ allele that contribute to the molecular characterization of Q. This induced mutation cause an Ala→Thr substitution at a highly conserved position in the first of two APETALA2 like domains of the Q protein. The mutation correlates with a speltoid phenotype as expected for a loss of Q function. Thus, it is demonstrated that the first APETALA2 like domain is essential for Q function. The phenotype was discovered in a mutant from an in silico TILLING population and the causative mutation could be identified in the existing sequence data. This report therefore provides an example of a forward genetic application of in silico TILLING.

Dominance and G×E interaction effects improve genomic prediction and genetic gain in intermediate wheatgrass (Thinopyrum intermedium).

Genomic selection (GS) based recurrent selection methods were developed to accelerate the domestication of intermediate wheatgrass [IWG, Thinopyrum intermedium (Host) Barkworth & D.R. Dewey]. A subset of the breeding population phenotyped at multiple environments is used to train GS models and then predict trait values of the breeding population. In this study, we implemented several GS models that investigated the use of additive and dominance effects and G×E interaction effects to understand how they affected trait predictions in intermediate wheatgrass. We evaluated 451 genotypes from the University of Minnesota IWG breeding program for nine agronomic and domestication traits at two Minnesota locations during 2017-2018. Genet-mean based heritabilities for these traits ranged from 0.34 to 0.77. Using four-fold cross validation, we observed the highest predictive abilities (correlation of 0.67) in models that considered G×E effects. When G×E effects were fitted in GS models, trait predictions improved by 18%, 15%, 20%, and 23% for yield, spike weight, spike length, and free threshing, respectively. Genomic selection models with dominance effects showed only modest increases of up to 3% and were trait-dependent. Cross-environment predictions were better for high heritability traits such as spike length, shatter resistance, free threshing, grain weight, and seed length than traits with low heritability and large environmental variance such as spike weight, grain yield, and seed width. Our results confirm that GS can accelerate IWG domestication by increasing genetic gain per breeding cycle and assist in selection of genotypes with promise of better performance in diverse environments.

The soft glumes of common wheat are sterile-lemmas as determined by the domestication gene Q

The Q gene in common wheat encodes an APETALA2 (AP2) transcription factor that causes the free threshing attribute. Wheat spikelets bearing several florets are subtended by a pair of soft glumes that allow free liberation of seeds. In wild species, the glumes are tough and rigid, making threshing difficult. However, the nature of these “soft glumes”, caused by the domestication allele Q is not clear. Here, we found that over expression of Q in common wheat leads to homeotic florets at glume positions. We provide phenotypic, microscopy, and marker genes evidence to demonstrate that the soft glumes of common wheat are in fact lemma-like organs, or so-called sterile-lemmas. By comparing the structures subtending spikelets in wheat and other crops such as rice and maize, we found that AP2 genes may play conserved functions in grasses by manipulating vestigial structures, such as floret-derived soft glumes in wheat and empty glumes in rice. Conversion of these seemingly vegetative organs to reproductive organs may be useful in yield improvement of crop species.

Breeding opportunities for early, free-threshing and semi-dwarf Triticum monococcum L.

Einkorn wheat, Triticum monococcum L. (2n = 2x = 14, AmAm genome), is a primitive, cultivated form of diploid wheat. The shortcoming of einkorn is that it lacks the free-threshing habit. Early heading and semi-dwarf traits are also required to fit modern agricultural practice. In the present study we developed T. monococcum pre-breeding germplasm having early, free threshing traits by utilizing an early heading source, two sources of soft glume (spike) and three sources of semi-dwarfism to combine their phenotypes into pre-breeding materials. We found two different genes determined free threshing of einkorn wheat. One of them was the sog (soft glume) gene from Triticum sinskajae Filat. et Kurkiev (2n = 2x = 14, AmAm genome) and another was the sos (soft spike) gene, which was completely linked or pleiotropic with the gene for semi-dwarfism. The genes sos, spd (short peduncle) and sd17654 (semi-dwarf CItr 17654) were utilized to develop semi-dwarf T. monococcum lines. Field performance of 6 early and free-threshing pre-breeding materials with sos and spd genes were tested over three crop seasons. Five semi-dwarf pre-breeding materials (PBMs) were obtained. However, these materials had slightly less grain yield than #252 (tall and hulled check) and PBM-1 (tall free-threshing check). Harvest index of the pre-breeding materials was improved due to the presence of sos and spd genes. If optimized cultivation practice is performed, these pre-breeding materials can be utilized as sources of early, free-threshing and semi-dwarf traits to produce modern T. monococcum varieties.

New alleles of the wheat domestication geneQreveal multiple roles in growth and reproductive development

The advantages of free threshing in wheat led to the selection of the domesticated Q allele, which is now present in almost all modern wheat varieties. Q and the pre-domestication allele, q, encode an AP2 transcription factor, with the domesticated allele conferring a free-threshing character and a subcompact (i.e. partially compact) inflorescence (spike). We demonstrate that mutations in the miR172 binding site of the Q gene are sufficient to increase transcript levels via a reduction in miRNA-dependent degradation, consistent with the conclusion that a single nucleotide polymorphism in the miRNA binding site of Q relative to q was essential in defining the modern Q allele. We describe novel gain- and loss-of-function alleles of Q and use these to define new roles for this gene in spike development. Q is required for the suppression of 'sham ramification', and increased Q expression can lead to the formation of ectopic florets and spikelets (specialized inflorescence branches that bear florets and grains), resulting in a deviation from the canonical spike and spikelet structures of domesticated wheat.

Breeding Value of Primary Synthetic Wheat Genotypes for Grain Yield.

To introduce new genetic diversity into the bread wheat gene pool from its progenitor, Aegilops tauschii (Coss.) Schmalh, 33 primary synthetic hexaploid wheat genotypes (SYN) were crossed to 20 spring bread wheat (BW) cultivars at the International Wheat and Maize Improvement Center. Modified single seed descent was used to develop 97 populations with 50 individuals per population using first back-cross, biparental, and three-way crosses. Individuals from each cross were selected for short stature, early heading, flowering and maturity, minimal lodging, and free threshing. Yield trials were conducted under irrigated, drought, and heat-stress conditions from 2011 to 2014 in Ciudad Obregon, Mexico. Genomic estimated breeding values (GEBVs) of parents and synthetic derived lines (SDLs) were estimated using a genomic best linear unbiased prediction (GBLUP) model with markers in each trial. In each environment, there were SDLs that had higher GEBVs than their recurrent BW parent for yield. The GEBVs of BW parents for yield ranged from -0.32 in heat to 1.40 in irrigated trials. The range of the SYN parent GEBVs for yield was from -2.69 in the irrigated to 0.26 in the heat trials and were mostly negative across environments. The contribution of the SYN parents to improved grain yield of the SDLs was highest under heat stress, with an average GEBV for the top 10% of the SDLs of 0.55 while the weighted average GEBV of their corresponding recurrent BW parents was 0.26. Using the pedigree-based model, the accuracy of genomic prediction for yield was 0.42, 0.43, and 0.49 in the drought, heat and irrigated trials, respectively, while for the marker-based model these values were 0.43, 0.44, and 0.55. The SYN parents introduced novel diversity into the wheat gene pool. Higher GEBVs of progenies were due to introgression and retention of some positive alleles from SYN parents.

Neolithic cultivation of tetraploid free threshing wheat in Denmark and Northern Germany: implications for crop diversity and societal dynamics of the Funnel Beaker Culture

For the first time, rachis fragments of tetraploid free threshing wheat were discovered for the Early Neolithic of the North European Plain, proving its cultivation by farmers of the Neolithic Funnel Beaker North group. A compilation of finds of free threshing wheat for Northern Europe based on a temporal resolution of Funnel Beaker sub-periods shows the importance of free threshing wheat—independent of genomic constitution—in the Early Neolithic Funnel Beaker crop assemblage. Based on this new evidence, we assume that the crop assemblages of the first farmers on the North European Plain consisted of three main cereals: emmer, barley and free threshing wheat. Both the new finds and the compilation of free threshing wheat finds support the scenario of a chronological separation of different agricultural systems that were implemented within different Funnel Beaker phases. Furthermore, the evidence for tetraploid free threshing wheat shows that the tradition of early Funnel Beaker groups is linked to southwestern agricultural developments, for example, of the Michelsberg group.

Over expression of the wheat BEL1-like gene TaqSH1 affects floral organ abscission in Arabidopsis thaliana

Seed shattering is one of the major domestication traits of crops. In wheat, except for the Q gene whose mutation renders free threshing and changing of rachis fragility, not much is known about the molecular mechanism for this process. We report here the cloning and characterization of TaqSH1, the ortholog of the rice seed shattering gene qSH1. TaqSH1 encodes a BEL1-like protein that is conserved between monocots and eudicots. TaqSH1 was located on the homoeologous group 3, a potential new genetic locus for seed threshability in wheat. Over expression of TaqSH1 in Arabidopsis resulted in dwarfed plants. The inflorescences of transgenic plants were more compact with larger pedicel angles. Scanning Electron Microscopy (SEM) showed that the transgenic siliques had a narrower replum where the dehiscence zone was altered. In addition, abscission of petals was significantly delayed due to delayed abscission zone development. Real-time PCR assays showed that over expression of TaqSH1 down regulated known Arabidopsis abscission related genes IDA, HAESA, KNAT1/6 and SHP1/2 in the transgenic plants. Taken together, our data suggest that TaqSH1 may represent another example of conserved mechanisms across monocots and eudicots for fruit/grain abscission and should have potential application in genetic manipulation of wheat seed shattering.

Evolution of tetraploid wheat based on variations in 5′ UTR regions of Ppd-A1: evidence of gene flow between emmer and timopheevi wheat

Previous study showed that tetraploid wheat was divided into two groups (Type AI and Type AII) based on sequences around Ppd-A1 gene (Takenaka and Kawahara in Theor Appl Genet 125(5):999–1014, 2012). That study focused on domesticated emmer wheat and used only 19 wild emmer wheats, so could not be clear the evolutional relationship between Type AI and Type AII. Here, a total of 669 accessions comprising 65 einkorn wheats, 185 wild emmer wheats, 107 hulled emmer wheats, 204 free-threshing (FT) emmer wheats, and 108 timopheevii wheats were studied by PCR assay and DNA sequencing for Type AI/AII. Type AII was an older type than Type AI because all einkorn accessions had Type AII. In wild emmer, Type AI was distributed in the northeast regions of its distribution and Type AII was found to be centered on Israel. A total of 37.4 % of hulled emmer accessions were Type AI, while 92.2 % of FT emmer accessions were Type AI. Differences in the proportion of Type AI/AII in domesticated emmer suggested a strong bottle-neck effect. We also found two MITE-like sequence deletion patterns from a part of Type AII accessions (dic-del and ara-del). Dic-del was found from only Israeli wild emmer accessions and ara-del was found from almost all timopheevii wheat accessions. Only three timopheevii accessions did not have ara-del, and one wild emmer accession and ten hulled emmer accessions had ara-del. These accessions suggested gene flow between emmer and timopheevii wheat.

Evolution and dispersal of emmer wheat (Triticum sp.) from novel haplotypes of Ppd-1 (photoperiod response) genes and their surrounding DNA sequences

The sequence data from 5' UTR, intronic, coding and 3' UTR regions of Ppd-A1 and Ppd-B1 were investigated for a total of 158 accessions of emmer wheat landraces comprising 19 of wild emmer wheat (Triticum dicoccoides), 45 of hulled emmer wheat (T. dicoccum) and 94 of free-threshing (FT) emmer wheat (T. durum etc.). We detected some novel types of deletions in the coding regions from 22 hulled emmer accessions and 20 FT emmer accessions. Emmer wheat accessions with these deletions could produce predicted proteins likely to lack function. We also observed some novel mutations in Ppd-B1. Sixty-seven and forty-one haplotypes were found in Ppd-A1 and Ppd-B1, respectively. Some mutations found in this study have not been known, so they have potential for useful genetic resources for wheat breeding. On the basis of sequence data from the 5' UTR region, both Ppd-A1 and Ppd-B1 haplotypes were divided into two groups (Type AI/AII and Type BI/BII). Types AI and AII of Ppd-A1 suggested gene flow between wild and hulled emmer. On the other hand, Types BI and BII of Ppd-B1 suggested gene flow between wild and FT emmer. More than half of hulled emmer accessions were Type AII/BI but few FT emmer accessions were of this type. Therefore, over half of the hulled emmer did not contribute to evolution of FT emmer.

Plant stores at pottery Neolithic Höyücek, southwest Turkey

AbstractEleven samples comprising an estimated 39,000 plant remains were analysed from a burnt destruction level at the pottery Neolithic site of Höyücek, southwest Turkey (radiocarbon dated 7550–7350 uncalibrated bp, 6400–6100 calibrated BC). Large stores of emmer (Triticum dicoccum), free threshing wheat (Triticum aestivum/durum), lentils (Lens culinaris), bitter vetch (Vicia ervilia) and chickpea (Cicer arietinum) were identified and these plants were interpreted as crops. The low levels of weeds and crop processing by-products suggest most of the samples were remains of stores of human food. Two samples in which wild components (for example,Triticum boeticum, Medicago, Aegilops) dominated were interpreted as crop processing by-products, presumably stored for fodder. The presence of these stores in a structure interpreted as having a religious function shows that domestic activities also took place there. Comparison with other Neolithic and Chalcolithic sites of west central Turkey demonstrates a good correspondence in the range of crops. The poor representation of barley at Höyücek doubtless reflects the small number of samples from the site.

Inheritance of threshability in synthetic hexaploid (Triticum turgidum×T. tauschii) by T. aestivum crosses

Abstract Triticum tauschii provides breeders with a valuable source of resistance and tolerance genes. Elucidation of the inheritance of traits in this species that hinder its use in breeding programmes is therefore of interest to wheat breeders. Inheritance of threshability was investigated in the crosses of four non‐free‐threshing (NFT) synthetic hexaploids (Triticum turgidum×T. tauschii) and two free‐threshing (FT) T. aestivum cultivars during four crop seasons over 3 years at E1 Batan and Ciudad Obregon, Mexico. The parents, their F1 Hybrids and individual F2 plant‐derived F3 progenies of the crosses revealed that ‘Altar 84’/T. tauschii (219), ‘Chen’/T. tauschii (205), ‘Chen’/T. tauschii (224), and ‘Duergand’/T. tauschii (214) have independently segregating loci with two dominant alleles controlling threshability. Intercrosses among the synthetics, except ‘Altar 84’/T. tauschii (219), showed the genes to be allelic to each other. The cross between the FT cultivars showed no segregation in the F3 generation, indicating common recessive genes. Based on these findings, population sizes of the synthetic‐derived breeding materials should be increased to improve the chances of selecting FT desirable plants in the programme.

Effect of Artificial Selection on Grain Dormancy in Eleusine (Gramineae)

The effect of artificial selection on grain dormancy was studied in six populations belonging to three species of Eleusine. Selection was effective in all pop- ulations but to different degrees. Genes for non-dormancy approached fixation at the fourth generation in one population of E. indica. Threshed grains exhibited higher degrees of germination in all selection cycles when compared with non- threshed grains. This resulted from the presence of inhibitors in the lemma and palea. Populations of E. coracana subsp. africana underwent an occasional drop in germination. This was attributed to a concomitant selection of ill-adapted genes or to fluctuation in allele frequency. It was concluded that man probably eliminated dormancy in finger-millet under domestication within a relatively short time period, particularly in cultivars used for malting. In response to selection by man, cereals and other seed crops have undergone well-defined evolutionary changes during their domestica- tion. In general, these changes were toward larger inflorescences, larger seeds, higher seed number, more determinate growth, free threshing, loss of seed dormancy, and others (Harlan et al. 1973; de Wet 1975). These evolutionary trends have not yet been well-documented experi- mentally. In this paper the efficiency of artificial selection on seed dor- mancy was examined in order to assess the dimensions of this aspect of cereal domestication. MATERIALS AND METHODS Selection against dormancy was applied to three populations of Eleu- sine indica (L.) Gaertn., two of E. coracana subsp. africana (Kennedy- O'Byrne) Hilu & de Wet, and one of E. tristachya (Lam.) Lam. These species are annual wild relatives of the African cereal Eleusine coracana subsp. coracana (L.) Gaertn. (finger millet). They were used in this study because 1) Eleusine species have a short life cycle that is insensitive to day length and a high seed set per individual plant due to extensive tillering, 2) populations of the wild progenitor of finger millet, E. coracana subsp. africana, are extant, and 3) the crop is used for both brewing and bread- making and requires immediate seed germination after harvesting to produce malt. These taxa are predominantly self-pollinated with 1-3% outcrossing. Plant materials were obtained from wild stands (table 1) by Professors J. R. Harlan and J. M. J. de Wet of the University of Illinois. Two 500-grain samples were taken from a mixed collection of each pop- ulation. The size of this initial sample was determined by low frequency of non-dormant seeds in natural populations, the average seed set per individual plant, and our intention to establish population densities com-

SEED PROTEIN PROFILES AND THE ORIGIN OF THE HEXAPLOID WHEATS

Protein profiles of Triticum and Aegilops species were obtained by electrophoresis of crude seed extracts on polyacrylamide gels. All subspecies of the hexaploid T. aestivum (AABBDD) showed a very uniform profile that could be closely simulated only by the pattern produced by a protein mixture (2:1) from specific profile types of the ancient tetraploid cultivar T. dicoccum (AABB) and the wild diploid Ae. squarrosa (DD). An exceptional hexaploid pattern occurred only in some accessions of T. aestivum ssp. macha. These results confirm the parentage of the aestivum hexaploids in general as T. dicoccum and Ae. squarrosa and more specifically identify the type of the D‐genome donor. They suggest that these wheats, excepting the aberrant macha types, had essentially a monophyletic origin in southwestern Asia. They favor the hypotheses that the cultivated aestivum wheats were derived from the so‐called primitive spelta complex primarily by mutation of a single gene governing the free threshing character and that alpine spelta represents an element displaced from the area of endemism.

Seed Protein Profiles and the Origin of the Hexaploid Wheats

Protein profiles of Triticum and Aegilops species were obtained by electrophoresis of crude seed extracts on polyacrylamide gels. All subspecies of the hexaploid T. aestivum (AABBDD) showed a very uniform profile that could be closely simulated only by the pattern produced by a protein mixture (2:1) from specific profile types of the ancient tetraploid cultivar T. dicoccum (AABB) and the wild diploid Ae. squarrosa (DD). An exceptional hexaploid pattern occurred only in some accessions of T. aestivum ssp. macha. These results confirm the parentage of the aestivum hexaploids in general as T. dicoccum and Ae. squarrosa and more specifically identify the type of the D-genome donor. They suggest that these wheats, excepting the aberrant macha types, had essentially a monophyletic origin in southwestern Asia. They favor the hypotheses that the cultivated aestivum wheats were derived from the so-called primitive spelta complex primarily by mutation of a single gene governing the free threshing character and that alpine spelta represents an element displaced from the area of endemism.