Intermediate wheatgrass (Thinopyrum intermedium; 2n = 6x = 42; IWG) is a perennial grass and is genetically related to common wheat, belonging to the Triticeae tribe of Poaideae (Mahelka et al., 2011). IWG produces large biomass and is among the most productive cool-season forage species in the western United States (Harmoney, 2015). As a perennial species, IWG provides substantial environmental services relative to annual grain crops, including reduced soil and water erosion, reduced soil nitrate leaching, increased carbon sequestration, and reduced input of seed, tillage, energy and pesticides (Culman et al., 2013; Glover et al., 2010; Robertson et al., 2000). IWG has a more extensive root system which can capture more applied fertilizer and reduce total nitrate leaching by 86% or more relative to annual wheat (Culman et al., 2013).
The direct domestication of IWG was initiated by Rodale Research Center, Kutztown, PA, in 1983 (Figure 1). IWG was identified as the most promising perennial grain crop among nearly 100 species of perennial grasses. IWG produced seeds with the thousand-grain weight as 5.3 grams on average, and the seeds can be mechanically harvested and threshed (Wagoner, 1990). The nutritional qualities of IWG are similar to wheat, but IWG has higher protein level and higher content of the sulfur containing amino acids, and whole flour of IWG grain performed well in baked products (Wagoner, 1990). After two cycles of selection performed at the Big Flats Plant Materials Center (Corning, NY), USDA Natural Resources Conservation Service, for grain yield and seed quality, the selected best plants were passed to scientists at the Land Institute, Salina, KS (Figure 1). Since 2003, scientists at The Land Institute (TLI) have been working on the domestication of IWG by selection for improved yield per head, increased seed size, free threshability, reduced height, and early maturity. After two cycles of selection, the grain yield was increased by about 77% and seed size by about 23%, when IWG were grown in solid-seed plots (DeHaan et al., 2014). In 2011, the University of Minnesota (St. Paul, USA), joined the domestication effort with the germplam from the third cycle of selection supplied by TLI (Figure 1). The primary goal of our IWG breeding program is to develop varieties with high grain yield while producing ecosystem services.
Figure 1. The history of intermediate wheatgrass domestication for grain yield. Recurrent selection was performed to improve yield per head, seed size, threshability, semi-dwarf stature, and early maturity. At the University of Minnesota, the breeding germplasm were derived from 66 female genets in the 3rd selection cycle of The Land Institute (TLI-C3).
The basic breeding strategy is pedigree and phenotype-based recurrent selection, i.e., selecting best plants from the best families (DeHaan et al., 2014). From 2011 to now, we finished two selection cycles. The seed size, grain yield, threshability and shattering resistance were greatly improved (Figure 2). The mean of the seed size in the breeding population was increased from 6.90mg to 10.90mg. The best genets (individual plants) produced seeds up to 17.3mg, half of the seed size of common wheat. This phenotype-based selection, however, is labor and time intensive as a result of planting, weeding, harvesting, and trait measurements on a few thousand spaced plants.
Figure 2. The progress in the improvement of IWG on seed size and seed shattering.
Plant breeding is being revolutionized by DNA marker technologies. The cost of markers is being dramatically reduced by next-generation sequencing. These marker technologies can be used with any species, even those species like IWG without previous genomics resources. Using genotyping-by-sequencing, we discovered genome-wide markers. And using 3,883 genome-wide markers discovered in 1,126 representative genets, we developed genomic prediction models. High predictive ability was observed for biomass, seed weight, plant height, grain yield, threshability and heading date using cross validation and independent validation, ranging from 0.46 for biomass to 0.67 for seed weight. Thus, we proposed a genomic selection-based breeding scheme (Figure 3), and have designed experiments to verify whether this breeding scheme provides faster genetic advance than the conventional breeding method.
Figure 3. Genomic selection-based breeding scheme. The genomic selection models trained using a subset of an IWG breeding population are used to estimate the breeding values of the remaining of the breeding population. A subset of the breeding population with 10 genets from each family is planted in the field. The genotypic and phenotypic data will be collected to develop genomic selection models. Based on the phenotypic data, the best families will be determined. Genets of the best families will be genotyped, and their breeding value will be estimated using the genomic selection models. The best plants will be selected and crossed in the green house. Their seeds will be used for the next selection cycle. The triangles, circles, squares and diamonds stand for genets from different families.
We optimized the genotyping-by-sequencing method to discover genome-wide markers in IWG. Using genotyping-by-sequencing, we identified 3,436 markers from a full-sib population and established the first genetic map of IWG with 21 linkage groups corresponding to 21 chromosomes. The IWG linkage groups showed high synteny and collinearity with barley as three homoeologous IWG linkage groups corresponded to one barley chromosome (Figure 4).
Figure 4. The collinearity of linkage groups of intermediate wheatgrass with barley chromosomes. The linkage groups were designed as LG1 to LG21, with LG1-3 corresponding to 1H, LG4-6 to 2H, and so on. The Spearman rank correlation coefficients (ρ) were calculated using R package ‘pspearman’. ** indicates significance at p < 0.01.
Current work was also conducted to identify the major quantitative trait loci (QTL) related to the agronomic traits using linkage mapping and association mapping. Within five years, we will be able to make crosses using parents possessing superior QTL, and pyramid the superior QTLs to improve IWG germplasm for disease resistance and agronomic traits. QTL identification and application will greatly accelerate the breeding process of IWG and help to improve commercial IWG cultivars.
Further efforts, collaborating with The Land Institute and Kansas State University, are focused on developing a draft genome assembly for IWG, along with generating reference transcriptome data sets that will be used for annotating gene models of the genome assembly, developing ultra-high-density genetic maps from genotyping by sequencing, and anchoring/ordering genomic scaffolds using population sequencing (POPSEQ). In light of the quickly evolving DNA sequencing tools and related technologies for optical mapping and long-range scaffolding information [cell-free Hi-C for assembly and genome organization (Chicago), 10X Genomics)], the initial draft assemblies developed from these efforts will improve as technologies mature.
James A. Anderson, Professor, Department of Agronomy and Plant Genetics
Kevin Betts, Senior Scientist, Department of Agronomy and Plant Genetics
Lee DeHaan, Plant Breeder, The Land Institute, Salina, Kansas
Kevin Dorn, Post Doctoral Research Associate, Kansas State University
Chathurada Suge Gajadeera, Post Doctoral Research Associate, Department of Food Science and Nutrition
Pam Ismail, Associate Professor, Department of Food Science and Nutrition
Jacob Jungers, Researcher, Department of Agronomy and Plant Genetics
Linda Kinkel, Professor, Department of Plant Pathology
Amy Mathiowetz, Research Assistant, Department of Food Science and Nutrition
Gary Muehlbauer, Professor, Department of Agronomy and Plant Genetics
Helene Murray, Adjunct Assistant Professor, Department of Agronomy and Plant Genetics
Devin Peterson, Professor, Department of Food Science and Nutrition
Jesse Poland, Assistant Professor, Kansas State University
Citra Rahardjo, Research Assistant, Department of Food Science and Nutrition
Tonya Schoenfuss, Associate Professor, Department of Food Science and Nutrition
Craig Sheaffer, Professor, Department of Agronomy and Plant Genetics
Catrin Tyl, Post-Doctoral Researcher, Department of Agronomy and Plant Genetics
M. Scott Wells, Assistant Professor, Department of Agronomy and Plant Genetics
Don Wyse, Professor, Department of Agronomy and Plant Genetics
Xiaofei Zhang, Researcher, Department of Agronomy and Plant Genetics
Initiative for Renewable Energy & the Environment
Forever Green Initiative
The Land Institute
Kansas State University
USDA-ARS, Cereal Crops Research Unit, Hard Spring and Durum Wheat Quality Laboratory,
USDA-ARS, Forage & Range Research Lab
Minnesota Crop Improvement Association