Rebecca J. McGee

Genetic diversity, population structure and genome-wide marker-trait association analysis emphasizing seed nutrients of the USDA pea (Pisum sativum L.) core collection

Genetic diversity, population structure and genome-wide marker-trait association analysis was conducted for the USDA pea (Pisum sativum L.) core collection. The core collection contained 285 accessions with diverse phenotypes and geographic origins. The 137 DNA markers included 102 polymorphic fragments amplified by 15 microsatellite primer pairs, 36 RAPD loci and one SCAR (sequence characterized amplified region) marker. The 49 phenotypic traits fall into the categories of seed macro- and micro-nutrients, disease resistance, agronomic traits and seed characteristics. Genetic diversity, population structure and marker-trait association were analyzed with the software packages PowerMarker, STUCTURE and TASSEL, respectively. A great amount of variation was revealed by the DNA markers at the molecular level. Identified were three sub-populations that constituted 56.1%, 13.0% and 30.9%, respectively, of the USDA Pisum core collection. The first sub-population is comprised of all cultivated pea varieties and landraces; the second of wild P. sativum ssp. elatius and abyssinicum and the accessions from the Asian highland (Afghanistan, India, Pakistan, China and Nepal); while the third is an admixture containing alleles from the first and second sub-populations. This structure was achieved using a stringent cutoff point of 15% admixture (q-value 85%) of the collection. Significant marker-trait associations were identified among certain markers with eight mineral nutrient concentrations in seed and other important phenotypic traits. Fifteen pairs of associations were at the significant levels of P ≤ 0.01 when tested using the three statistical models. These markers will be useful in marker-assisted selection to breed pea cultivars with desirable agronomic traits and end-user qualities.

Lentils (Lens culinaris L.), a Rich Source of Folates

The potential for genetic biofortification of U.S.-grown lentils ( Lens culinaris L.) with bioavailable folate has not been widely studied. The objectives of this study were (1) to determine the folate concentration of 10 commercial lentil cultivars grown in Minot and McLean counties, North Dakota, USA, in 2010 and 2011, (2) to determine the genotype (G) × environmental (E) interactions for folate concentration in lentil cultivars, and (3) to compare the folate concentration of other pulses [field peas ( Pisum sativum L.) and chickpea ( Cicer arietinum L.)] grown in the United States. Folate concentration in lentil cultivars ranged from 216 to 290 μg/100 g with a mean of 255 μg/100 g. In addition, lentil showed higher folate concentration compared to chickpea (42-125 μg/100 g), yellow field pea (41-55 μg/100 g), and green field pea (50-202 μg/100 g). A 100 g serving of lentils could provide a significant amount of the recommended daily allowance of dietary folates (54-73%) for adults. A significant year × location interaction on lentil folate concentration was observed; this indicates that possible location sourcing may be required for future lentil folate research.

Genetic Diversity of Cultivated Lentil (Lens culinaris Medik.) and Its Relation to the World's Agro-ecological Zones

Assessment of genetic diversity and population structure of germplasm collections plays a critical role in supporting conservation and crop genetic enhancement strategies. We used a cultivated lentil (Lens culinaris Medik.) collection consisting of 352 accessions originating from 54 diverse countries to estimate genetic diversity and genetic structure using 1194 polymorphic single nucleotide polymorphism (SNP) markers which span the lentil genome. Using principal coordinate analysis, population structure analysis and UPGMA cluster analysis, the accessions were categorized into three major groups that prominently reflected geographical origin (worlds agro-ecological zones). The three clusters complemented the origins, pedigrees, and breeding histories of the germplasm. The three groups were (a) South Asia (sub-tropical savannah), (b) Mediterranean, and (c) northern temperate. Based on the results from this study, it is also clear that breeding programs still have considerable genetic diversity to mine within the cultivated lentil, as surveyed South Asian and Canadian germplasm revealed narrow genetic diversity.

Genome-wide association mapping of partial resistance to Aphanomyces euteiches in pea

BackgroundGenome-wide association (GWA) mapping has recently emerged as a valuable approach for refining the genetic basis of polygenic resistance to plant diseases, which are increasingly used in integrated strategies for durable crop protection. Aphanomyces euteiches is a soil-borne pathogen of pea and other legumes worldwide, which causes yield-damaging root rot. Linkage mapping studies reported quantitative trait loci (QTL) controlling resistance to A. euteiches in pea. However the confidence intervals (CIs) of these QTL remained large and were often linked to undesirable alleles, which limited their application in breeding. The aim of this study was to use a GWA approach to validate and refine CIs of the previously reported Aphanomyces resistance QTL, as well as identify new resistance loci.MethodsA pea-Aphanomyces collection of 175 pea lines, enriched in germplasm derived from previously studied resistant sources, was evaluated for resistance to A. euteiches in field infested nurseries in nine environments and with two strains in climatic chambers. The collection was genotyped using 13,204 SNPs from the recently developed GenoPea Infinium® BeadChip.ResultsGWA analysis detected a total of 52 QTL of small size-intervals associated with resistance to A. euteiches, using the recently developed Multi-Locus Mixed Model. The analysis validated six of the seven previously reported main Aphanomyces resistance QTL and detected novel resistance loci. It also provided marker haplotypes at 14 consistent QTL regions associated with increased resistance and highlighted accumulation of favourable haplotypes in the most resistant lines. Previous linkages between resistance alleles and undesired late-flowering alleles for dry pea breeding were mostly confirmed, but the linkage between loci controlling resistance and coloured flowers was broken due to the high resolution of the analysis. A high proportion of the putative candidate genes underlying resistance loci encoded stress-related proteins and others suggested that the QTL are involved in diverse functions.ConclusionThis study provides valuable markers, marker haplotypes and germplasm lines to increase levels of partial resistance to A. euteiches in pea breeding.

A community resource for exploring and utilizing genetic diversity in the USDA pea single plant plus collection

Globally, pea (Pisum sativum L.) is an important temperate legume crop for food, feed and fodder, and many breeding programs develop cultivars adapted to these end-uses. In order to assist pea development efforts, we assembled the USDA Pea Single Plant Plus Collection (PSPPC), which contains 431 P. sativum accessions with morphological, geographic and taxonomic diversity. The collection was characterized genetically in order to maximize its value for trait mapping and genomics-assisted breeding. To that end, we used genotyping-by-sequencing—a cost-effective method for de novo single-nucleotide polymorphism (SNP) marker discovery—to generate 66 591 high-quality SNPs. These data facilitated the identification of accessions divergent from mainstream breeding germplasm that could serve as sources of novel, favorable alleles. In particular, a group of accessions from Central Asia appear nearly as diverse as a sister species, P. fulvum, and subspecies, P. sativum subsp. elatius. PSPPC genotypes can be paired with new and existing phenotype data for trait mapping; as proof-of-concept, we localized Mendel’s A gene controlling flower color to its known position. We also used SNP data to define a smaller core collection of 108 accessions with similar levels of genetic diversity as the entire PSPPC, resulting in a smaller germplasm set for research screening and evaluation under limited resources. Taken together, the results presented in this study along with the release of a publicly available SNP data set comprise a valuable resource for supporting worldwide pea genetic improvement efforts.

Phenotyping Nutritional and Antinutritional Traits

Evolution of nutrient-rich food systems to calorie-focused production agriculture has created serious agricultural and human health issues: marginalization of traditional agricultural crops, greater dependence of agricultural inputs, and creation of both energy and micronutrient malnutrition. To date more than half of global human populations are suffering numerous health problems associated with excess calories and lack of essential micronutrients. Pulse crops, in particular lentils, are promising crops not only to improve human health but also to reduce agricultural inputs toward greater agricultural sustainability. In this book chapter, human micronutrient malnutrition issues, suggestions to reduce micronutrient deficiencies, promise of pulse crops using lentil as an example, lentil’s micronutrient and antinutrient profiles, nutrient analytical procedures, and the needs to shift our thinking from calorie-focused to nutrient-focused approaches are also presented.

Dietary Fiber Analysis of Four Pulses Using AOAC 2011.25: Implications for Human Health

Chickpeas, common beans, dry peas, and lentils are pulse crops that have been a cornerstone of the human diet since the inception of agriculture. However, the displacement of pulses from the diet by low fiber protein alternatives has resulted in a pervasive deficiency referred to as the dietary fiber gap. Using an analytical method American Association of Analytical Chemists (AOAC) 2011.25 that conforms to the Codex Alimentarius Commission consensus definition for dietary fiber, the fiber content of these pulse crops was evaluated in seed types used for commercial production. These pulse crops have 2 to 3 times more fiber per 100 g edible portion than other dietary staples. Moreover, there is marked variation in fiber content among cultivars of the same crop. We conclude that pulse crop consumption should be emphasized in efforts to close the dietary fiber gap. The substantial differences in fiber content among currently available cultivars within a crop can be used to further improve gains in fiber intake without the need to change dietary habits. This provides a rationale for cultivar-based food labeling.

Development and validation of breeder-friendly KASPar markers for er1 , a powdery mildew resistance gene in pea ( Pisum sativum L.)

Powdery mildew of pea is caused by Erysiphe pisi DC and is a serious threat to pea (Pisum sativum L.) production throughout much of the world. Development and utilization of genetic resistance to powdery mildew is considered an effective and sustainable strategy to manage this disease. One gene, er1, conferring powdery mildew resistance, was previously cloned and sequenced, and the functional markers for each resistance allele were reported. Allele-specific DNA markers are efficient and powerful tools to facilitate crop improvement and new cultivar development in breeding programs. However, extensive application of these markers is limited by gel-associated obstacles. In this study, eight breeder-friendly kompetitive allele-specific PCR (KASPar) markers were developed to overcome the problems of gel-based markers and increase the efficiency of genotypic screening. In order to identify additional pea germplasm with powdery mildew resistance, these KASPar markers were deployed and used to genotype a pea collection derived from the USDA pea single-plant (PSP) collection. Simultaneously, a phenotypic screening and a genotypic validation using the corresponding gel-based functional markers were conducted on the PSP collection. One pea accession, PI 142775, was identified by both phenotyping and genotyping to carry the allele er1-1 for powdery mildew resistance, indicating that the KASPar assay is an efficient and robust tool for breeding for powdery mildew resistance.

Near-infrared spectroscopic models for analysis of winter pea (Pisum sativum L.) quality constituents: Near-infrared spectroscopic models for analysis of winter pea

BACKGROUND Winter pea (Pisum sativum L.) grows well in a wide geographic region, both as a forage and cover crop. Understanding the quality constituents of this crop is important for both end uses; however, analysis of quality constituents by conventional wet chemistry methods is laborious, slow and costly. Near infrared reflectance spectroscopy (NIRS) is a precise, accurate, rapid and cheap alternative to using wet chemistry for estimating quality constituents. We developed and validated NIRS calibration models for constituent analysis of this crop. RESULTS Of the 11 constituent models developed, nine constituents including moisture, dry-matter, total-nitrogen, crude protein, acid detergent fiber, neutral detergent fiber, AD-lignin, cellulose and non-fibrous carbohydrate had low standard errors and a high coefficient of determination (R2 = 0.88-0.98; 1 - VR, which is the coefficient of determination during cross-validation = 0.77-0.92) for both calibration and cross-validation, indicating their potential for quantitative predictability. The calibration models for ash (R2 = 0.65; 1 - VR = 0.46) and hemicellulose (R2 = 0.75; 1 - VR = 0.50) also appeared to be adequate for qualitative screening. Predictions of an independent validation set yielded reliable agreement between the NIRS predicted values and the reference values with low standard error of prediction (SEP), low bias, high coefficient of determination (r2 = 0.82-0.95), high ratios of performance to deviation (RPD = SD/SEP; 2.30-3.85) and high ratios of performance to interquartile distance (RPIQ = IQ/SEP; 2.57-7.59) for all 11 constituents. CONCLUSION Precise, accurate and rapid analysis of winter pea for major forage and cover crop quality constituents can be performed at a low cost using the NIRS calibration models developed.

Will selenium fertilization improve biological nitrogen fixation in lentils

ABSTRACT Lentil is a cool season food legume rich in protein and micronutrients. The objective of this study was to determine the effect of a low dosage of selenium (Se) on biological nitrogen (N) fixation, seed Se, and grain yield in lentils. The experiment was carried out at the Carrington Research and Extension Center, North Dakota, USA in 2012 and 2013. Six lentil genotypes were treated with three Se treatments. Application of selenate significantly increased percent Nderived from air (%Ndfa; 44%) compared to selenite (38%) and control (37%). In addition, selenate significantly increased lentil seed Se (1129 µg kg−1) compared to selenite (844 µg kg−1) and the control (542 µg kg−1). Both %Ndfa and grain yield increased with Se application. Selenate was the most effective form to increase %Ndfa. More research is required to determine the biochemical relationships between lentil yield and the Nfixation under Se deficient soils.