Ability of Genomic Prediction to Bi-Parent-Derived Breeding Population Using Public Data for Soybean Oil and Protein Content.

Genomic selection (GS) is a marker-based selection method used to improve the genetic gain of quantitative traits in plant breeding. A large number of breeding datasets are available in the soybean database, and the application of these public datasets in GS will improve breeding efficiency and reduce time and cost. However, the most important problem to be solved is how to improve the ability of across-population prediction. The objectives of this study were to perform genomic prediction (GP) and estimate the prediction ability (PA) for seed oil and protein contents in soybean using available public datasets to predict breeding populations in current, ongoing breeding programs. In this study, six public datasets of USDA GRIN soybean germplasm accessions with available phenotypic data of seed oil and protein contents from different experimental populations and their genotypic data of single-nucleotide polymorphisms (SNPs) were used to perform GP and to predict a bi-parent-derived breeding population in our experiment. The average PA was 0.55 and 0.50 for seed oil and protein contents within the bi-parents population according to the within-population prediction; and 0.45 for oil and 0.39 for protein content when the six USDA populations were combined and employed as training sets to predict the bi-parent-derived population. The results showed that four USDA-cultivated populations can be used as a training set individually or combined to predict oil and protein contents in GS when using 800 or more USDA germplasm accessions as a training set. The smaller the genetic distance between training population and testing population, the higher the PA. The PA increased as the population size increased. In across-population prediction, no significant difference was observed in PA for oil and protein content among different models. The PA increased as the SNP number increased until a marker set consisted of 10,000 SNPs. This study provides reasonable suggestions and methods for breeders to utilize public datasets for GS. It will aid breeders in developing GS-assisted breeding strategies to develop elite soybean cultivars with high oil and protein contents.

Genetic Dissection of Diverse Seed Coat Patterns in Cowpea through a Comprehensive GWAS Approach.

This study investigates the genetic determinants of seed coat color and pattern variations in cowpea (Vigna unguiculata), employing a genome-wide association approach. Analyzing a mapping panel of 296 cowpea varieties with 110,000 single nucleotide polymorphisms (SNPs), we focused on eight unique coat patterns: (1) Red and (2) Cream seed; (3) White and (4) Brown/Tan seed coat; (5) Pink, (6) Black, (7) Browneye and (8) Red/Brown Holstein. Across six GWAS models (GLM, SRM, MLM, MLMM, FarmCPU from GAPIT3, and TASSEL5), 13 significant SNP markers were identified and led to the discovery of 23 candidate genes. Among these, four specific genes may play a direct role in determining seed coat pigment. These findings lay a foundational basis for future breeding programs aimed at creating cowpea varieties aligned with consumer preferences and market requirements.

Low frequency of the wild-type freezing-tolerance LsCBF7 allele among lettuce population suggests a negative selection during domestication and breeding.

KEY MESSAGE: Sustainable winter production in lettuce requires freezing tolerant varieties. This study identified a wild-type allele of LsCBF7 that could contribute to freezing tolerance improvement in lettuce. Lettuce is one of the most consumed vegetables globally. While ideally grown in 13-21 °C, its cultivation extends into winter in milder climates. However, occasional freezing temperatures can significantly reduce yields. Therefore, the development of freezing-tolerant lettuce varieties has become a long-term goal of lettuce breeding programs. Despite its significance, our understanding of freezing tolerance in lettuce remains limited. Plants have evolved a coping mechanism against freezing, known as cold acclimation, whereby they can increase freezing tolerance when pre-exposed to low nonfreezing temperatures. The CBF pathway is well-known for its central role in cold acclimation. Previously, we identified 14 CBF genes in lettuce and discovered that one of them, LsCBF7, had a loss-of-function mutation. In this study, we uncovered that accessions from colder regions carried the wild-type allele of LsCBF7 and this allele likely contributed to increased freezing tolerance, with 14% of the lettuce population carrying this allele. Interestingly, in wild lettuce (L. serriola) that is considered a progenitor of cultivated lettuce, this wild-type allele was much more common, with a frequency of 90%. This finding suggests that this wild-type allele may have undergone negative selection during the domestication or breeding of lettuce. Our data strongly indicate that this allele could be linked to early bolting, an undesirable trait in lettuce, which may have driven the negative selection. While this wild-type allele shows promise for improving freezing tolerance in lettuce, it is crucial to decouple it from the early bolting trait to fully harness its potential in lettuce breeding.

Evaluation of Tomato Germplasm against Tomato Brown Rugose Fruit Virus and Identification of Resistance in Solanum pimpinellifolium.

The tomato is one of the most important vegetable crops grown worldwide. Tomato brown rugose fruit virus (ToBRFV), a seed-borne tobamovirus, poses a serious threat to tomato production due to its ability to break the resistant genes (Tm-1, Tm-2, Tm-22) in tomatoes. The objective of this work was to identify new resistant source(s) of tomato germplasm against ToBRFV. To achieve this aim, a total of 476 accessions from 12 Solanum species were tested with the ToBRFV US isolate for their resistance and susceptibility. As a result, a total of 44 asymptomatic accessions were identified as resistant/tolerant, including thirty-one accessions of S. pimpinellifolium, one accession of S. corneliomulleri, four accessions of S. habrochaites, three accessions of S. peruvianum, and five accessions of S. subsection lycopersicon hybrid. Further analyses using serological tests identified four highly resistant S. pimpinellifolium lines, PI 390713, PI 390714, PI 390716, and PI 390717. The inheritance of resistance in the selected lines was verified in the next generation and confirmed using RT-qPCR. To our knowledge, this is a first report of high resistance to ToBRFV in S. pimpinellifolium. These new genetic resources will expand the genetic pool available for breeders to develop new resistant cultivars of tomato against ToBRFV.

Genome-Wide Association Study Reveals a QTL Region for Ashy Stem Blight Resistance in PRA154 Andean Common Bean.

Ashy stem blight (ASB) caused by Macrophomina phaseolina (Tassi) Goidanich affects the common bean (Phaseolus vulgaris L.) at all growing stages. Higher levels of resistance were observed in Andean common beans, but specific resistant quantitative trait loci (QTLs) conferring resistance to this pathogen have not been reported in this gene pool. The objectives of this research were to: (i) conduct a genome-wide association study (GWAS) and QTL mapping for resistance in the Andean breeding line PRA154; and (ii) identify single nucleotide polymorphism (SNP) markers and candidate genes for ASB resistance. Phenotyping was conducted under greenhouse conditions by inoculating the 107 F6:7 recombinant inbred lines (RILs) derived from the cross between the susceptible cultivar 'Verano' and the partial-resistant breeding line PRA154 twice with the M. phaseolina isolate PRI21. Genotyping was performed with 109,040 SNPs distributed across all 11 P. vulgaris chromosomes. A novel major QTL was located between 28,761,668 and 31,263,845 bp, extending 2.5 Mbp on chromosome Pv07, and the highest significant SNP markers were Chr07_28761668_A_G, Chr07_29131720_G_A, and Chr07_31263845_C_T with the highest LOD (more than 10 in most of the cases) and R-squared values, explaining 40% of the phenotypic variance of the PRI21 isolate. The gene Phvul.007G173900 (methylcrotonyl-CoA carboxylase alpha chain and mitochondrial 3-methylcrotonyl-CoA carboxylase 1 [MCCA]) with a size of 10,891 bp, located between 29,131,591 and 29,142,481 bp on Pv07, was identified as one candidate for ASB resistance in PRA154, and it contained Chr07_29131720_G_A. The QTL and genetic marker information could be used to assist common bean breeders to develop germplasm and cultivars with ASB resistance through molecular breeding.

Genome-wide characterization and evolutionary analysis of the AP2/ERF gene family in lettuce (Lactuca sativa).

The APETALA2/ETHYLENE RESPONSIVE FACTOR (AP2/ERF) gene family plays vital roles in plants, serving as a key regulator in responses to abiotic stresses. Despite its significance, a comprehensive understanding of this family in lettuce remains incomplete. In this study, we performed a genome-wide search for the AP2/ERF family in lettuce and identified a total of 224 members. The duplication patterns provided evidence that both tandem and segmental duplications contributed to the expansion of this family. Ka/Ks ratio analysis demonstrated that, following duplication events, the genes have been subjected to purifying selection pressure, leading to selective constraints on their protein sequence. This selective pressure provides a dosage benefit against stresses in plants. Additionally, a transcriptome analysis indicated that some duplicated genes gained novel functions, emphasizing the contribution of both dosage effect and functional divergence to the family functionalities. Furthermore, an orthologous relationship study showed that 60% of genes descended from a common ancestor of Rosid and Asterid lineages, 28% from the Asterid ancestor, and 12% evolved in the lettuce lineage, suggesting lineage-specific roles in adaptive evolution. These results provide valuable insights into the evolutionary mechanisms of the AP2/ERF gene family in lettuce, with implications for enhancing abiotic stress tolerance, ultimately contributing to the genetic improvement of lettuce crop production.

Genetic Architecture of Salt Tolerance in Cowpea (Vigna unguiculata (L.) Walp.) at Seedling Stage Using a Whole Genome Resequencing Approach.

Cowpea (Vigna unguiculata (L.) Walp.) is a diploid legume crop used for human consumption, feed for livestock, and cover crops. Earlier reports have shown that salinity has been a growing threat to cowpea cultivation. The objectives of this study were to conduct a genome-wide association study (GWAS) to identify SNP markers and to investigate candidate genes for salt tolerance in cowpea. A total of 331 cowpea genotypes were evaluated for salt tolerance by supplying a solution of 200 mM NaCl in our previous work. The cowpea panel was genotyped using a whole genome resequencing approach, generating 14,465,516 SNPs. Moreover, 5,884,299 SNPs were used after SNP filtering. GWAS was conducted on a total of 296 cowpea genotypes that have high-quality SNPs. BLINK was used for conducting GWAS. Results showed (1) a strong GWAS peak on an 890-bk region of chromosome 2 for leaf SPAD chlorophyll under salt stress in cowpea and harboring a significant cluster of nicotinamide adenine dinucleotide (NAD) dependent epimerase/dehydratase genes such as Vigun02g128900.1, Vigun02g129000.1, Vigun02g129100.1, Vigun02g129200.1, and Vigun02g129500.1; (2) two GWAS peaks associated with relative tolerance index for chlorophyll were identified on chromosomes 1 and 2. The peak on chromosome 1 was defined by a cluster of 10 significant SNPs mapped on a 5 kb region and was located in the vicinity of Vigun01g086000.1, encoding for a GATA transcription factor. The GWAS peak on chromosome 2 was defined by a cluster of 53 significant SNPs and mapped on a 68 bk region of chromosome 2, and (3) the highest GWAS peak was identified on chromosome 3, and this locus was associated with leaf score injury. This peak was within the structure of a potassium channel gene (Vigun03g144700.1). To the best of our knowledge, this is one the earliest reports on the salt tolerance study of cowpea using whole genome resequencing data.

Genome-Wide Association Study and Genomic Prediction of Fusarium Wilt Resistance in Common Bean Core Collection.

The common bean (Phaseolus vulgaris L.) is a globally cultivated leguminous crop. Fusarium wilt (FW), caused by Fusarium oxysporum f. sp. phaseoli (Fop), is a significant disease leading to substantial yield loss in common beans. Disease-resistant cultivars are recommended to counteract this. The objective of this investigation was to identify single nucleotide polymorphism (SNP) markers associated with FW resistance and to pinpoint potential resistant common bean accessions within a core collection, utilizing a panel of 157 accessions through the Genome-wide association study (GWAS) approach with TASSEL 5 and GAPIT 3. Phenotypes for Fop race 1 and race 4 were matched with genotypic data from 4740 SNPs of BARCBean6K_3 Infinium Bea Chips. After ranking the 157-accession panel and revealing 21 Fusarium wilt-resistant accessions, the GWAS pinpointed 16 SNPs on chromosomes Pv04, Pv05, Pv07, Pv8, and Pv09 linked to Fop race 1 resistance, 23 SNPs on chromosomes Pv03, Pv04, Pv05, Pv07, Pv09, Pv10, and Pv11 associated with Fop race 4 resistance, and 7 SNPs on chromosomes Pv04 and Pv09 correlated with both Fop race 1 and race 4 resistances. Furthermore, within a 30 kb flanking region of these associated SNPs, a total of 17 candidate genes were identified. Some of these genes were annotated as classical disease resistance protein/enzymes, including NB-ARC domain proteins, Leucine-rich repeat protein kinase family proteins, zinc finger family proteins, P-loopcontaining nucleoside triphosphate hydrolase superfamily, etc. Genomic prediction (GP) accuracy for Fop race resistances ranged from 0.26 to 0.55. This study advanced common bean genetic enhancement through marker-assisted selection (MAS) and genomic selection (GS) strategies, paving the way for improved Fop resistance.

A Genome-Wide Association Study Reveals Region Associated with Seed Protein Content in Cowpea.

Cowpea (Vigna unguiculata L. Walp., 2n = 2x = 22) is a protein-rich crop that complements staple cereals for humans and serves as fodder for livestock. It is widely grown in Africa and other developing countries as the primary source of protein in the diet; therefore, it is necessary to identify the protein-related loci to improve cowpea breeding. In the current study, we conducted a genome-wide association study (GWAS) on 161 cowpea accessions (151 USDA germplasm plus 10 Arkansas breeding lines) with a wide range of seed protein contents (21.8~28.9%) with 110,155 high-quality whole-genome single-nucleotide polymorphisms (SNPs) to identify markers associated with protein content, then performed genomic prediction (GP) for future breeding. A total of seven significant SNP markers were identified using five GWAS models (single-marker regression (SMR), the general linear model (GLM), Mixed Linear Model (MLM), Fixed and Random Model Circulating Probability Unification (FarmCPU), and Bayesian-information and Linkage-disequilibrium Iteratively Nested Keyway (BLINK), which are located at the same locus on chromosome 8 for seed protein content. This locus was associated with the gene Vigun08g039200, which was annotated as the protein of the thioredoxin superfamily, playing a critical function for protein content increase and nutritional quality improvement. In this study, a genomic prediction (GP) approach was employed to assess the accuracy of predicting seed protein content in cowpea. The GP was conducted using cross-prediction with five models, namely ridge regression best linear unbiased prediction (rrBLUP), Bayesian ridge regression (BRR), Bayesian A (BA), Bayesian B (BB), and Bayesian least absolute shrinkage and selection operator (BL), applied to seven random whole genome marker sets with different densities (10 k, 5 k, 2 k, 1 k, 500, 200, and 7), as well as significant markers identified through GWAS. The accuracies of the GP varied between 42.9% and 52.1% across the seven SNPs considered, depending on the model used. These findings not only have the potential to expedite the breeding cycle through early prediction of individual performance prior to phenotyping, but also offer practical implications for cowpea breeding programs striving to enhance seed protein content and nutritional quality.

Skim resequencing finely maps the downy mildew resistance loci RPF2 and RPF3 in spinach cultivars whale and Lazio.

Commercial production of spinach (Spinacia oleracea L.) is centered in California and Arizona in the US, where downy mildew caused by Peronospora effusa is the most destructive disease. Nineteen typical races of P. effusa have been reported to infect spinach, with 16 identified after 1990. The regular appearance of new pathogen races breaks the resistance gene introgressed in spinach. We attempted to map and delineate the RPF2 locus at a finer resolution, identify linked single nucleotide polymorphism (SNP) markers, and report candidate downy mildew resistance (R) genes. Progeny populations segregating for RPF2 locus derived from resistant differential cultivar Lazio were infected using race 5 of P. effusa and were used to study for genetic transmission and mapping analysis in this study. Association analysis performed with low coverage whole genome resequencing-generated SNP markers mapped the RPF2 locus between 0.47 to 1.46 Mb of chromosome 3 with peak SNP (Chr3_1, 221, 009) showing a LOD value of 61.6 in the GLM model in TASSEL, which was within 1.08 Kb from Spo12821, a gene that encodes CC-NBS-LRR plant disease resistance protein. In addition, a combined analysis of progeny panels of Lazio and Whale segregating for RPF2 and RPF3 loci delineated the resistance section in chromosome 3 between 1.18-1.23 and 1.75-1.76 Mb. This study provides valuable information on the RPF2 resistance region in the spinach cultivar Lazio compared to RPF3 loci in the cultivar Whale. The RPF2 and RPF3 specific SNP markers, plus the resistant genes reported here, could add value to breeding efforts to develop downy mildew resistant cultivars in the future.

A genome-wide association study and genomic prediction for Phakopsora pachyrhizi resistance in soybean.

Soybean brown rust (SBR), caused by Phakopsora pachyrhizi, is a devastating fungal disease that threatens global soybean production. This study conducted a genome-wide association study (GWAS) with seven models on a panel of 3,082 soybean accessions to identify the markers associated with SBR resistance by 30,314 high quality single nucleotide polymorphism (SNPs). Then five genomic selection (GS) models, including Ridge regression best linear unbiased predictor (rrBLUP), Genomic best linear unbiased predictor (gBLUP), Bayesian least absolute shrinkage and selection operator (Bayesian LASSO), Random Forest (RF), and Support vector machines (SVM), were used to predict breeding values of SBR resistance using whole genome SNP sets and GWAS-based marker sets. Four SNPs, namely Gm18_57,223,391 (LOD = 2.69), Gm16_29,491,946 (LOD = 3.86), Gm06_45,035,185 (LOD = 4.74), and Gm18_51,994,200 (LOD = 3.60), were located near the reported P. pachyrhizi R genes, Rpp1, Rpp2, Rpp3, and Rpp4, respectively. Other significant SNPs, including Gm02_7,235,181 (LOD = 7.91), Gm02_7234594 (LOD = 7.61), Gm03_38,913,029 (LOD = 6.85), Gm04_46,003,059 (LOD = 6.03), Gm09_1,951,644 (LOD = 10.07), Gm10_39,142,024 (LOD = 7.12), Gm12_28,136,735 (LOD = 7.03), Gm13_16,350,701(LOD = 5.63), Gm14_6,185,611 (LOD = 5.51), and Gm19_44,734,953 (LOD = 6.02), were associated with abundant disease resistance genes, such as Glyma.02G084100, Glyma.03G175300, Glyma.04g189500, Glyma.09G023800, Glyma.12G160400, Glyma.13G064500, Glyma.14g073300, and Glyma.19G190200. The annotations of these genes included but not limited to: LRR class gene, cytochrome 450, cell wall structure, RCC1, NAC, ABC transporter, F-box domain, etc. The GWAS based markers showed more accuracies in genomic prediction than the whole genome SNPs, and Bayesian LASSO model was the ideal model in SBR resistance prediction with 44.5% ~ 60.4% accuracies. This study aids breeders in predicting selection accuracy of complex traits such as disease resistance and can shorten the soybean breeding cycle by the identified markers.

A Genome-Wide Association Study and Genomic Prediction for Fiber and Sucrose Contents in a Mapping Population of LCP 85-384 Sugarcane.

Sugarcane (Saccharum spp. hybrids) is an economically important crop for both sugar and biofuel industries. Fiber and sucrose contents are the two most critical quantitative traits in sugarcane breeding that require multiple-year and multiple-location evaluations. Marker-assisted selection (MAS) could significantly reduce the time and cost of developing new sugarcane varieties. The objectives of this study were to conduct a genome-wide association study (GWAS) to identify DNA markers associated with fiber and sucrose contents and to perform genomic prediction (GP) for the two traits. Fiber and sucrose data were collected from 237 self-pollinated progenies of LCP 85-384, the most popular Louisiana sugarcane cultivar from 1999 to 2007. The GWAS was performed using 1310 polymorphic DNA marker alleles with three models of TASSEL 5, single marker regression (SMR), general linear model (GLM) and mixed linear model (MLM), and the fixed and random model circulating probability unification (FarmCPU) of R package. The results showed that 13 and 9 markers were associated with fiber and sucrose contents, respectively. The GP was performed by cross-prediction with five models, ridge regression best linear unbiased prediction (rrBLUP), Bayesian ridge regression (BRR), Bayesian A (BA), Bayesian B (BB) and Bayesian least absolute shrinkage and selection operator (BL). The accuracy of GP varied from 55.8% to 58.9% for fiber content and 54.6% to 57.2% for sucrose content. Upon validation, these markers can be applied in MAS and genomic selection (GS) to select superior sugarcane with good fiber and high sucrose contents.

Resequencing worldwide spinach germplasm for identification of field resistance QTLs to downy mildew and assessment of genomic selection methods.

Downy mildew, commercially the most important disease of spinach, is caused by the obligate oomycete Peronospora effusa. In the past two decades, new pathogen races have repeatedly overcome the resistance used in newly released cultivars, urging the need for more durable resistance. Commercial spinach cultivars are bred with major R genes to impart resistance to downy mildew pathogens and are effective against some pathogen races/isolates. This work aimed to evaluate the worldwide USDA spinach germplasm collections and commercial cultivars for resistance to downy mildew pathogen in the field condition under natural inoculum pressure and conduct genome wide association analysis (GWAS) to identify resistance-associated genomic regions (alleles). Another objective was to evaluate the prediction accuracy (PA) using several genomic prediction (GP) methods to assess the potential implementation of genomic selection (GS) to improve spinach breeding for resistance to downy mildew pathogen. More than four hundred diverse spinach genotypes comprising USDA germplasm accessions and commercial cultivars were evaluated for resistance to downy mildew pathogen between 2017-2019 in Salinas Valley, California and Yuma, Arizona. GWAS was performed using single nucleotide polymorphism (SNP) markers identified via whole genome resequencing (WGR) in GAPIT and TASSEL programs; detected 14, 12, 5, and 10 significantly associated SNP markers with the resistance from four tested environments, respectively; and the QTL alleles were detected at the previously reported region of chromosome 3 in three of the four experiments. In parallel, PA was assessed using six GP models and seven unique marker datasets for field resistance to downy mildew pathogen across four tested environments. The results suggest the suitability of GS to improve field resistance to downy mildew pathogen. The QTL, SNP markers, and PA estimates provide new information in spinach breeding to select resistant plants and breeding lines through marker-assisted selection (MAS) and GS, eventually helping to accumulate beneficial alleles for durable disease resistance.

Identification of a QTL region for ashy stem blight resistance using genome-wide association and linage analysis in common bean recombinant inbred lines derived from BAT 477 and NY6020-4.

Ashy stem blight (ASB), caused by the fungus Macrophomina phaseolina (Tassi) Goidanich is an important disease of the common bean (Phaseolus vulgaris L.). It is important to identify quantitative trait loci (QTL) for ASB resistance and introgress into susceptible cultivars of the common bean. The objective of this research was to identify QTL and single nucleotide polymorphism (SNP) markers associated with ASB resistance in recombinant inbred lines (RIL) derived from a cross between BAT 477 and NY6020-4 common bean. One hundred and twenty-six F6:7 RIL were phenotyped for ASB in the greenhouse. Disease severity was scored on a scale of 1-9. Genotyping was performed using whole genome resequencing with 2x common bean genome size coverage, and over six million SNPs were obtained. After being filtered, 72,017 SNPs distributed on 11 chromosomes were used to conduct the genome-wide association study (GWAS) and QTL mapping. A novel QTL region of ~4.28 Mbp from 35,546,329 bp to 39,826,434 bp on chromosome Pv03 was identified for ASB resistance. The two SNPs, Chr03_39824257 and Chr03_39824268 located at 39,824,257 bp and 39,824,268 bp on Pv03, respectively, were identified as the strongest markers associated with ASB resistance. The gene Phvul.003G175900 (drought sensitive, WD repeat-containing protein 76) located at 39,822,021 - 39,824,655 bp on Pv03 was recognized as one candidate for ASB resistance in the RIL, and the gene contained the two SNP markers. QTL and SNP markers may be used to select plants and lines for ASB resistance through marker-assisted selection (MAS) in common bean breeding.

Mapping and selection of downy mildew resistance in spinach cv. whale by low coverage whole genome sequencing.

Spinach (Spinacia oleracea) is a popular leafy vegetable crop and commercial production is centered in California and Arizona in the US. The oomycete Peronospora effusa causes the most important disease in spinach, downy mildew. A total of nineteen races of P. effusa are known, with more than 15 documented in the last three decades, and the regular emergence of new races is continually overcoming the genetic resistance to the pathogen. This study aimed to finely map the downy mildew resistance locus RPF3 in spinach, identify single nucleotide polymorphism (SNP) markers associated with the resistance, refine the candidate genes responsible for the resistance, and evaluate the prediction performance using multiple machine learning genomic prediction (GP) methods. Segregating progeny population developed from a cross of resistant cultivar Whale and susceptible cultivar Viroflay to race 5 of P. effusa was inoculated under greenhouse conditions to determine downy mildew disease response across the panel. The progeny panel and the parents were resequenced at low coverage (1x) to identify genome wide SNP markers. Association analysis was performed using disease response phenotype data and SNP markers in TASSEL, GAPIT, and GENESIS programs and mapped the race 5 resistance loci (RPF3) to 1.25 and 2.73 Mb of Monoe-Viroflay chromosome 3 with the associated SNP in the 1.25 Mb region was 0.9 Kb from the NBS-LRR gene SOV3g001250. The RPF3 locus in the 1.22-1.23 Mb region of Sp75 chromosome 3 is 2.41-3.65 Kb from the gene Spo12821 annotated as NBS-LRR disease resistance protein. This study extended our understanding of the genetic basis of downy mildew resistance in spinach cultivar Whale and mapped the RPF3 resistance loci close to the NBS-LRR gene providing a target to pursue functional validation. Three SNP markers efficiently selected resistance based on multiple genomic selection (GS) models. The results from this study have added new genomic resources, generated an informed basis of the RPF3 locus resistant to spinach downy mildew pathogen, and developed markers and prediction methods to select resistant lines.

Growth and Photosynthetic Responses of Cowpea Genotypes under Waterlogging at the Reproductive Stage.

Waterlogging is an important environmental stress limiting the productivity of crops worldwide. Cowpea (Vigna unguiculata L.) is particularly sensitive to waterlogging stress during the reproductive stage, with a consequent decline in pod formation and yield. However, little is known about the critical processes underlying cowpea's responses to waterlogging during the reproductive stage. Thus, we investigated the key parameters influencing carbon fixation, including stomatal conductance (gs), intercellular CO2 concentration, chlorophyll content, and chlorophyll fluorescence, of two cowpea genotypes with contrasting waterlogging tolerance. These closely related genotypes have starkly contrasting responses to waterlogging during and after 7 days of waterlogging stress (DOW). In the intolerant genotype ('EpicSelect.4'), waterlogging resulted in a gradual loss of pigment and decreased photosynthetic capacity as a consequent decline in shoot biomass. On the other hand, the waterlogging-tolerant genotype ('UCR 369') maintained CO2 assimilation rate (A), stomatal conductance (gs), biomass, and chlorophyll content until 5 DOW. Moreover, there was a highly specific downregulation of the mesophyll conductance (gm), maximum rate of Rubisco (Vcmax), and photosynthetic electron transport rate (Jmax) as non-stomatal limiting factors decreasing A in EpicSelect.4. Exposure of EpicSelect.4 to 2 DOW resulted in the loss of PSII photochemistry by downregulating the PSII quantum yield (Fv/Fm), photochemical efficiency (ΦPSII), and photochemical quenching (qP). In contrast, we found no substantial change in the photosynthesis and chlorophyll fluorescence of UCR 369 in the first 5 DOW. Instead, UCR 369 maintained biomass accumulation, chlorophyll content, and Rubisco activity, enabling the genotype to maintain nutrient absorption and photosynthesis during the early period of waterlogging. However, compared to the control, both cowpea genotypes could not fully recover their photosynthetic capacity after 7 DOW, with a more significant decline in EpicSelect.4. Overall, our findings suggest that the tolerant UCR 369 genotype maintains higher photosynthesis under waterlogging stress attributable to higher photochemical efficiency, Rubisco activity, and less stomatal restriction. After recovery, the incomplete recovery of A can be attributed to the reduced gs caused by severe waterlogging damage in both genotypes. Thus, promoting the rapid recovery of stomata from waterlogging stress may be crucial for the complete restoration of carbon fixation in cowpeas during the reproductive stage.

Waterlogging during the reproductive growth stage causes physiological and biochemical modifications in the leaves of cowpea (Vigna unguiculata L.) genotypes with contrasting tolerance.

Waterlogging causes various metabolic, physiological, and morphological changes in crops, resulting in yield loss of most legumes in rainfed and irrigated agriculture. However, research on cowpea genotypes using physiological and biochemical traits as a measure of tolerance to waterlogging stress is limited. We evaluated the impacts of 7 days of waterlogging (DOW) and 7 days of recovery (DOR) on the physiology and biochemistry of two cowpea (Vigna unguiculata (L.) Walp) genotypes (UCR 369 and EpicSelect.4) with contrasting waterlogging tolerance. Cowpea genotypes were grown in a controlled environment until the R2 stage and then subjected to 7 DOW. Later, the waterlogged plants were reoxygenated for an additional 7 DOR. Overall, cowpea genotypes had a contrasting response to waterlogging using different mechanisms. Compared to the control, the photosynthetic parameters of both cowpea genotypes were impaired under 7 DOW and could not recover at 7 DOR, with a larger decline in EpicSelect.4.7 DOW caused significant loss in the chlorophyll and carotenoid content of both genotypes. However, only waterlogged UCR 369 was not photo-inhibited and able to restore the levels of chlorophyll and carotenoids at 7 DOR. In addition, 7 DOW induced intense stress in UCR 369 with increased zeaxanthin, sucrose, and flavonoid content, while these metabolites were decreased in EpicSelect.4. On the other hand, glucose, fructose, and phenolic content were increased in EpicSelect.4 but decreased in UCR 369 at 7 DOR. In summary, compared to EpicSelect.4, UCR 369 restored their photosynthetic pigments and metabolites to the control levels at 7 DOR, indicating a likely tolerance to waterlogging stress.

Genetic dissection of nitrogen induced changes in the shoot and root biomass of spinach.

Efficient partitioning of above and below-ground biomass in response to nitrogen (N) is critical to the productivity of plants under sub-optimal conditions. It is particularly essential in vegetable crops like spinach with shallow root systems, a short growth cycle, and poor nitrogen use efficiency. In this study, we conducted a genome-wide association study (GWAS) to explore N-induced changes using spinach accessions with diverse genetic backgrounds. We evaluated phenotypic variations as percent changes in the shoot and root biomass in response to N using 201 spinach accessions grown in randomized complete blocks design in a soilless media under a controlled environment. A GWAS was performed for the percent changes in the shoot and root biomass in response to N in the 201 spinach accessions using 60,940 whole-genome resequencing generated SNPs. Three SNP markers, chr4_28292655, chr6_1531056, and chr6_37966006 on chromosomes 4 and 6, were significantly associated with %change in root weight, and two SNP markers, chr2_18480277 and chr4_47598760 on chromosomes 2 and 4, were significantly associated with % change shoot weight. The outcome of this study established a foundation for genetic studies needed to improve the partitioning of total biomass and provided a resource to identify molecular markers to enhance N uptake via marker-assisted selection or genomic selection in spinach breeding programs.

Identification of Candidate Genes and Genomic Selection for Seed Protein in Soybean Breeding Pipeline.

Soybean is a primary meal protein for human consumption, poultry, and livestock feed. In this study, quantitative trait locus (QTL) controlling protein content was explored via genome-wide association studies (GWAS) and linkage mapping approaches based on 284 soybean accessions and 180 recombinant inbred lines (RILs), respectively, which were evaluated for protein content for 4 years. A total of 22 single nucleotide polymorphisms (SNPs) associated with protein content were detected using mixed linear model (MLM) and general linear model (GLM) methods in Tassel and 5 QTLs using Bayesian interval mapping (IM), single-trait multiple interval mapping (SMIM), single-trait composite interval mapping maximum likelihood estimation (SMLE), and single marker regression (SMR) models in Q-Gene and IciMapping. Major QTLs were detected on chromosomes 6 and 20 in both populations. The new QTL genomic region on chromosome 6 (Chr6_18844283-19315351) included 7 candidate genes and the Hap.X AA at the Chr6_19172961 position was associated with high protein content. Genomic selection (GS) of protein content was performed using Bayesian Lasso (BL) and ridge regression best linear unbiased prediction (rrBULP) based on all the SNPs and the SNPs significantly associated with protein content resulted from GWAS. The results showed that BL and rrBLUP performed similarly; GS accuracy was dependent on the SNP set and training population size. GS efficiency was higher for the SNPs derived from GWAS than random SNPs and reached a plateau when the number of markers was >2,000. The SNP markers identified in this study and other information were essential in establishing an efficient marker-assisted selection (MAS) and GS pipelines for improving soybean protein content.