Breeding Tools Research Articles

Apomictic Malus plants exhibit abnormal pollen development.

Apomixis is the asexual reproduction through seeds that leads to the production of genetically uniform progeny. It has become an important tool in plant breeding because it facilitates the retention of genotypes with desirable traits and allows seeds to be obtained directly from mother plants. Apomixis is rare in most economically important crops, but it occurs in some Malus species. Here, the apomictic characteristics of Malus were examined using four apomictic and two sexually reproducing Malus plants. Results from transcriptome analysis showed that plant hormone signal transduction was the main factor affecting apomictic reproductive development. Four of the apomictic Malus plants examined were triploid, and pollen was either absent or present in very low densities in the stamen. Variation in the presence of pollen was associated with variation in the apomictic percentage; specifically, pollen was absent in the stamens of tea crabapple plants with the highest apomictic percentage. Furthermore, pollen mother cells failed to progress normally into meiosis and pollen mitosis, a trait mostly observed in apomictic Malus plants. The expression levels of meiosis-related genes were upregulated in apomictic plants. Our findings indicate that our simple method of detecting pollen abortion could be used to identify apple plants that are capable of apomictic reproduction.

Breeding high‐protein pigeonpea genotypes and their agronomic and biological assessments

AbstractProteins, inevitable for nutritional security of human beings and legumes, by far, are the cheapest source of this vital nutrient. The escalating prices and never halting population growth limit the per capita availability of protein‐rich legumes. In view of limited land resource and need to grow other food crops, the greater protein harvests are possible only by increasing the protein levels of popularly grown legumes. In this context, attempts were made for raising the protein content in pigeonpea [Cajanus cajan(L.) Millsp.] through traditional plant breeding tools. For this, the high‐protein trait was successfully transferred from wild relatives of pigeonpea to the cultivated types. In the derived inbred lines, the protein content was significantly enhanced from 20% ‐ 22% to 28% ‐ 30%. Two high‐protein lines HPL 40 and HPL 8 also produced 2100 and 1660 kg/ha grain yield, respectively. This simply means that, in comparison with traditional cultivars, the cultivation of high‐protein lines will provide additional 100 kg/ha of digestible protein to the farming family. This paper, besides describing the breeding procedures, also discusses the accomplishments of this breeding endeavour with respect to its various nutritional and biological properties.

Molecular Markers in Plant Breeding

Molecular markers are an important tool for plant breeding. Since the 1980s, in response to the technology development, molecular marker approaches have been further diversified. The establishment of new-generation sequencing and high-throughput plant phenotyping has greatly decreased the time to genotype large numbers of individuals. For breeders who are not very familiar with molecular techniques and want to catch up with the advances in the field, this review offers basic knowledge. Each molecular marker technology has specific advantages as well as limitations. Molecular marker types, diversity studies, QTL mapping, associative mapping, marker-assisted backcrossing and genomic selection are explored. Marker application in plant breeding is also described. In the genome, molecular markers can detect the genetic architecture of a trait, but also identify candidate genes with an important role in plant breeding programs.

Mastering DNA Content Estimation by Flow Cytometry as an Efficient Tool for Plant Breeding and Biodiversity Research.

Flow cytometry gives a unique opportunity to analyze thousands of individual cells for multiple parameters in a course of minutes. The most commonly used flow cytometry application in plant biology is estimation of nuclear DNA content. This becomes an indispensable tool in different areas of plant research, including breeding, taxonomy, plant development, evolutionary biology, populational studies and others. DNA content analysis can provide an insight into natural ploidy changes that reflect evolutionary processes, such as interspecific hybridization and polyploidization. It is also widely used for processing samples with biotechnologically induced ploidy changes, for instance, plants produced by doubled haploid technology. Absolute genome size data produced by cytometric analysis serve as useful taxon-specific markers since genome size vary between different taxa. It often allows the distinguishing of species within a genus or even different subspecies. Introducing flow cytometry method in the lab is extremely appealing, but new users face a significant challenge of learning instrument management, quality sample preparation and data processing. Not only is flow cytometry a complex method, but plant samples have unique features that make plants a demanding research subject. Without proper training, researchers risk damaging the expensive instrument or publishing poor quality data, artifacts or unreproducible results. We bring together information from our experience, key papers and online resources to provide step by step protocols and give a starting point for exploring the abundant cytometry literature.

Mechanistic Understanding of Leakage and Consequences and Recent Technological Advances in Improving Nitrogen Use Efficiency in Cereals

Although nitrogen (N) is the most limiting nutrient for agricultural production, its overuse is associated with environmental pollution, increased concentration of greenhouse gases, and several human and animal health implications. These implications are greatly affected by biochemical transformations and losses of N such as volatilization, leaching, runoff, and denitrification. Half of the globally produced N fertilizers are used to grow three major cereals—rice, wheat, and maize—and their current level of N recovery is approximately 30–50%. The continuously increasing application of N fertilizers, despite lower recovery of cereals, can further intensify the environmental and health implications of leftover N. To address these implications, the improvement in N use efficiency (NUE) by adopting efficient agronomic practices and modern breeding and biotechnological tools for developing N efficient cultivars requires immediate attention. Conventional and marker-assisted selection methods can be used to map quantitative trait loci, and their introgression in elite germplasm leads to the creation of cultivars with better NUE. Moreover, gene-editing technology gives the opportunity to develop high-yielding cultivars with improved N utilization capacity. The most reliable and cheap methods include agronomic practices such as site-specific N management, enhanced use efficiency fertilizers, resource conservation practices, precision farming, and nano-fertilizers that can help farmers to reduce the environmental losses of N from the soil–plant system, thus improving NUE. Our review illuminates insights into recent advances in local and scientific soil and crop management technologies, along with conventional and modern breeding technologies on how to increase NUE that can help reduce linked N pollution and health implications.

Towards Marker-Assisted Breeding for Black Rot Bunch Resistance: Identification of a Major QTL in the Grapevine Cultivar 'Merzling'.

Black rot (BR), caused by Guignardia bidwellii, is an emergent fungal disease threatening viticulture and affecting several mildew-tolerant varieties. However, its genetic bases are not fully dissected yet. For this purpose, a segregating population derived from the cross 'Merzling' (hybrid, resistant) × 'Teroldego' (V. vinifera, susceptible) was evaluated for BR resistance at the shoot and bunch level. The progeny was genotyped with the GrapeReSeq Illumina 20K SNPchip, and 7175 SNPs were combined with 194 SSRs to generate a high-density linkage map of 1677 cM. The QTL analysis based on shoot trials confirmed the previously identified Resistance to Guignardia bidwellii (Rgb)1 locus on chromosome 14, which explained up to 29.2% of the phenotypic variance, reducing the genomic interval from 2.4 to 0.7 Mb. Upstream of Rgb1, this study revealed a new QTL explaining up to 79.9% of the variance for bunch resistance, designated Rgb3. The physical region encompassing the two QTLs does not underlie annotated resistance (R)-genes. The Rgb1 locus resulted enriched in genes belonging to phloem dynamics and mitochondrial proton transfer, while Rgb3 presented a cluster of pathogenesis-related Germin-like protein genes, promoters of the programmed cell death. These outcomes suggest a strong involvement of mitochondrial oxidative burst and phloem occlusion in BR resistance mechanisms and provide new molecular tools for grapevine marker-assisted breeding.

Syntenic Cell Wall QTLs as Versatile Breeding Tools: Intraspecific Allelic Variability and Predictability of Biomass Quality Loci in Target Plant Species

Syntenic cell wall QTLs (SQTLs) can identify genetic determinants of biomass traits in understudied species based on results from model crops. However, their effective use in plant breeding requires SQTLs to display intraspecific allelic variability and to predict causative loci in other populations/species than the ones used for SQTLs identification. In this study, genome assemblies from different accessions of Arabidopsis, rapeseed, tomato, rice, Brachypodium and maize were used to evaluate the intraspecific variability of SQTLs. In parallel, a genome-wide association study (GWAS) on cell wall quality traits was performed in miscanthus to verify the colocalization between GWAS loci and miscanthus SQTLs. Finally, an analogous approach was applied on a set of switchgrass cell wall QTLs retrieved from the literature. These analyses revealed large SQTLs intraspecific genetic variability, ranging from presence-absence gene variation to SNPs/INDELs and changes in coded proteins. Cell wall genes displaying gene dosage regulation, such as PAL and CAD, displayed presence-absence variation in Brachypodium and rapeseed, while protein INDELs were detected for the Brachypodium homologs of the rice brittle culm-like 8 locus, which may likely impact cell wall quality. Furthermore, SQTLs significantly colocalized with the miscanthus and switchgrass QTLs, with relevant cell wall genes being retained in colocalizing regions. Overall, SQTLs are useful tools to screen germplasm for relevant genes and alleles to improve biomass quality and can increase the efficiency of plant breeding in understudied biomass crops.

Parametric mapping of QTL for resistance to European canker in apple in ‘Aroma’ × ‘Discovery’

Resistance to European canker (Neonectria ditissima) in apple is currently one of the most important breeding targets for commercial production in Sweden. Previous research has identified significant genetic variation in susceptibility to the disease, with the local Swedish cultivar ‘Aroma’ considered as one of the most resistant cultivars. Identification of genetic regions underlying the resistance of this cultivar would be a valuable tool for future breeding. Thus, we performed Bayesian quantitative trait loci (QTL) mapping for resistance to European canker in a full-sib family of ‘Aroma’ × ‘Discovery’. Mapping was performed with the area under the disease progression curves (AUDPCs) from all seven (AUDPC_All7) and the first four assessments (AUDPC_First4), and three parameters of a sigmoid growth model for lesion length. As a scale for the effect of the different parameters, historic phenotypic data from screenings of a genetically diverse germplasm was compiled and re-analyzed. The parametrization of the data on lesion growth increased the number of QTL that could be identified with high statistical power, and provided some insight into their roles during different stages of disease development in the current experimental setup. Five QTL regions with strong or decisive evidence were identified on linkage groups 1, 8, 15, and 16. The QTL regions could be assigned to either of the parameters lesion length at the first assessment (‘LL_A1’), the maximal lesion growth rate (lesion length doubling time, ‘t_gen’), and the lesion length at girdling (‘LL_G’). Three of these QTL were traced along the pedigrees of some known relatives of the FS family, and discussed in relation to future crosses for breeding and genetic research.

Complete chloroplast genome and phylogenetic analysis of Lonicera caerulea var. edulis (Caprifoliaceae)

Lonicera caerulea var. edulis, known as “blue honeysuckle” or “Haskap,” is a deciduous shrub that belongs to the Caprifoliaceae family. Characterized by the high cold hardiness and high quality of fruit, it has become a novel cash crop in cold regions worldwide. The lack of available chloroplast (cp) genome information limits studies of its molecular breeding and phylogeny. Here, the complete cp genome of Lonicera caerulea var. edulis was assembled and characterized for the first time. It was 155,142 bp in length with 38.43% of GC content, including 23,841 bp inverted repeat regions (IRs), an 88,737 bp large single-copy region (LSC), and an 18,723 bp small single-copy region (SSC). A total of 132 genes, including 85 protein-coding genes, 8 rRNA genes, and 39 tRNA genes were annotated. Phylogenetic analysis indicated that L. caerulea var. edulis was closely related to L. tangutica. These data and results provide a valuable resource for the development of breeding tools and genetic diversity studies for L. caerulea.

Phenotyping for the Seedling Stage of the Drought Stress Tolerance in Rice (Oryza sativa L.)

The ever-increasing demand for rice raises the need to increase productivity by developing drought-tolerant rice varieties. Drought tolerance is a complex polygenic trait that largely depends upon plant developmental stages and showed genotype-specific variability. The experiment was conducted using drought tolerant (Binadhan-19, BRRI dhan83) and drought susceptible (BRRI dhan26, BRRI dhan48) rice genotypes at the glasshouse of Bangladesh Institute of Nuclear Agriculture (BINA), Mymensingh to characterize the seedling stage drought tolerance as well as disclose the variability of the genotypes for morphological and biochemical traits. A completely randomized design (CRD) with three replications and treatments (control and drought stress imposed by 20% PEG) was used for the experiment. Data on shoot length, root length, fresh root weight, fresh shoot weight, dry root weight, dry shoot weight, chlorophyll content (SPAD value), MDA, and H2O2 were recorded and analyzed through appropriate breeding tools. Significant variation (p<0.01) was observed for genotype, treatment, and genotype (G) × treatment (T) interactions viz., shoot length, root length, root fresh weight, shoot fresh weight, root dry weight, shoot dry weight, chlorophyll, H2O2, and MDA. Drought stress leads to a significant decrease in root and shoots growth whereas the level of H2O2 and MDA increased significantly. A greater decrease in root and shoot growth was observed in susceptible genotypes (BRRI dhan28, BRRI dhan48) compared to tolerant genotypes (BRRI dhan71, Binadhan-19). In contrast, a limited increase in H2O2 and MDA was recorded in tolerant genotypes compared to susceptible genotypes. H2O2 showed a significant positive correlation with root and shoot characteristics under control conditions, whereas H2O2 showed a significant negative correlation with chlorophyll content under drought conditions. MDA showed a significant negative correlation with most of the studied traits under well-watered conditions. Considering all of the traits at the seedling stage, the genotype Binadhan-19 is considered a drought-tolerant genotype both under well-watered and drought conditions, and this genotype was selected for further study under direct field conditions as well as for genetic improvement against drought stress.

Genebanking plant genetic resources in the postgenomic era

Genebanking, the process of preserving genetic resources, is a central practice in the modern management of crop genetics, especially for the species used for food and agriculture. Closely interrelated networks of local, national and global actors are responsible for ex situ conservation. They all seek to make plant genetic resources accessible for all and now face new challenges arising from digitisation. Plant sciences are entering the postgenomic era, moving fast from initially providing a single reference genome for each species (genomics), to harnessing the extent of diversity within crop species (pangenomics) and among their relatives (referred to as postgenomics). This paper describes the extent to which ex situ collections have already undergone a digital shift, or are planning to do so, and the potential impact of this postgenomic-induced dematerialisation on the global governance of plant genetic resources. In turn, digitising material (seed) collection changes the relationship between genebanks and genomic databases. Comprehensive genomic characterisation of genebank accessions is ongoing, and I argue here that these efforts may provide a unique opportunity for genebanks to further embrace the moral, ethical and ultimately political principles on which they were built. Repurposing genebanks as decentralised digital biocentres could help relocate capabilities and stewardship over genetic resources. Empowering local farmers by providing access, promoting the use and unlocking benefits from state-of-the-art tools of modern plant breeding may allow bridging the breeding divide. However, to accomplish such a paradigm shift, genebanks require a strong political mandate that must primarily originate from the access and benefit-sharing framework. Only so may the global challenges associated with the loss of biodiversity and food insecurity be addressed.

Developing algae as a sustainable food source.

Current agricultural and food production practices are facing extreme stress, posed by climate change and an ever-increasing human population. The pressure to feed nearly 8 billion people while maintaining a minimal impact on the environment has prompted a movement toward new, more sustainable food sources. For thousands of years, both the macro (seaweed and kelp) and micro (unicellular) forms of algae have been cultivated as a food source. Algae have evolved to be highly efficient at resource utilization and have proven to be a viable source of nutritious biomass that could address many of the current food production issues. Particularly for microalgae, studies of their large-scale growth and cultivation come from the biofuel industry; however, this knowledge can be reasonably translated into the production of algae-based food products. The ability of algae to sequester CO2 lends to its sustainability by helping to reduce the carbon footprint of its production. Additionally, algae can be produced on non-arable land using non-potable water (including brackish or seawater), which allows them to complement rather than compete with traditional agriculture. Algae inherently have the desired qualities of a sustainable food source because they produce highly digestible proteins, lipids, and carbohydrates, and are rich in essential fatty acids, vitamins, and minerals. Although algae have yet to be fully domesticated as food sources, a variety of cultivation and breeding tools exist that can be built upon to allow for the increased productivity and enhanced nutritional and organoleptic qualities that will be required to bring algae to mainstream utilization. Here we will focus on microalgae and cyanobacteria to highlight the current advancements that will expand the variety of algae-based nutritional sources, as well as outline various challenges between current biomass production and large-scale economic algae production for the food market.

Mapping QTLs for Super-Earliness and Agro-Morphological Traits in RILs Population Derived from Interspecific Crosses between Pisum sativum × P. fulvum.

Earliness in crop plants has a vital role in prevention of heat-induced drought stress and in combating global warming, which is predicted to exacerbate in the near future. Furthermore, earliness may expand production into northern areas or higher altitudes, having relatively shorter growing season and may also expand arable lands to meet global food demands. The primary objective of the present study was to investigate quantitative trait loci (QTLs) for super-earliness and important agro-morphological traits in a recombinant inbred line (RIL) population derived from an interspecific cross. A population of 114 RILs developed through single-seed descent from an interspecific cross involving Pisum sativum L. and P. fulvum Sibth. et Sm. was evaluated to identify QTLs for super-earliness and important agro-morphological traits. A genetic map was constructed with 44 SSRs markers representing seven chromosomes with a total length of 262.6 cM. Of the 14 QTLs identified, two were for super-earliness on LG2, one for plant height on LG3, six for number of pods per plant on LG2, LG4, LG5 and LG6, one for number of seeds per pod on LG6, one for pod length on LG4 and three for harvest index on LG3, LG5, and LG6. AA205 and AA372-1 flanking markers for super-earliness QTLs were suggested for marker-assisted selection (MAS) in pea breeding programs due to high heritability of the trait. This is the first study to map QTLs originating from P. sativum and P. fulvum recently identified species with super-earliness character and the markers (AA205 and AA372-1) linked to QTLs were valuable molecular tools for pea breeding.

V-primer: software for the efficient design of genome-wide InDel and SNP markers from multi-sample variant call format (VCF) genotyping data.

DNA markers are indispensable tools in genetics and genomics research as well as in crop breeding, particularly for marker-assisted selection. Recent advances in next-generation sequencing technology have made it easier to obtain genome sequences for various crop species, enabling the large-scale identification of DNA polymorphisms among varieties, which in turn has made DNA marker design more accessible. However, existing primer design software is not suitable for designing many types of genome-wide DNA markers from next-generation sequencing data. Here, we describe the development of V-primer, high-throughput software for designing insertion/deletion, cleaved amplified polymorphic sequence, and single-nucleotide polymorphism (SNP) markers. We validated the applicability of these markers in different crops. In addition, we performed multiplex PCR targeted amplicon sequencing using SNP markers designed with V-primer. Our results demonstrate that V-primer facilitates the efficient and accurate design of primers and is thus a useful tool for genetics, genomics, and crop breeding. V-primer is freely available at https://github.com/ncod3/vprimer.

A Novel Intelligent System for Dynamic Observation of Cotton Verticillium Wilt.

Verticillium wilt is one of the most critical cotton diseases, which is widely distributed in cotton-producing countries. However, the conventional method of verticillium wilt investigation is still manual, which has the disadvantages of subjectivity and low efficiency. In this research, an intelligent vision-based system was proposed to dynamically observe cotton verticillium wilt with high accuracy and high throughput. Firstly, a 3-coordinate motion platform was designed with the movement range 6,100 mm × 950 mm × 500 mm, and a specific control unit was adopted to achieve accurate movement and automatic imaging. Secondly, the verticillium wilt recognition was established based on 6 deep learning models, in which the VarifocalNet (VFNet) model had the best performance with a mean average precision (mAP) of 0.932. Meanwhile, deformable convolution, deformable region of interest pooling, and soft non-maximum suppression optimization methods were adopted to improve VFNet, and the mAP of the VFNet-Improved model improved by 1.8%. The precision-recall curves showed that VFNet-Improved was superior to VFNet for each category and had a better improvement effect on the ill leaf category than fine leaf. The regression results showed that the system measurement based on VFNet-Improved achieved high consistency with manual measurements. Finally, the user software was designed based on VFNet-Improved, and the dynamic observation results proved that this system was able to accurately investigate cotton verticillium wilt and quantify the prevalence rate of different resistant varieties. In conclusion, this study has demonstrated a novel intelligent system for the dynamic observation of cotton verticillium wilt on the seedbed, which provides a feasible and effective tool for cotton breeding and disease resistance research.

Eliciting Targeted Mutations in Medicago sativa Using CRISPR/Cas9-Mediated Genome Editing: A Potential Tool for the Improvement of Disease Resistance.

CRISPR/Cas9 (clustered regularly interspaced short palindromic repeats/CRISPR-associated protein 9) has become a breeding tool of choice for eliciting targeted genetic alterations in crop species as a means of improving a wide range of agronomic traits, including disease resistance, in recent years. With the recent development of CRISPR/Cas9 technology in Medicago sativa (alfalfa), which is an important perennial forage legume grown worldwide, its use for the enhancement of pathogen resistance is almost certainly on the horizon. In this chapter, we present detailed procedures for the generation of a single nonhomologous end-joining-derived indel at a precise genomic locus of alfalfa via CRISPR/Cas9. This method encompasses crucial steps in this process, including guide RNA design, binary CRISPR vector construction, Agrobacterium-mediated transformation of alfalfa explants, and molecular assessments of transformed genotypes for transgene and edit identification.

Breeding and genomics approaches for improving phosphorus-use efficiency in grain legumes

Phosphorus (P) is an essential plant macronutrient, but P sources for plant growth are non-renewable, causing great concern for future sustainable agriculture and global food security. Thus, enhancing plant P-use efficiency (PUE) by improving P-acquisition and P-utilization efficiencies is urgently needed in various crops, including grain legumes, for intensive cropping systems. This review discusses how to harness the genetic variability in PUE traits across grain legume gene pools to improve PUE using various conventional breeding approaches and emerging breeding tools. The genetic architecture of PUE traits is complex, being quantitatively inherited and highly influenced by the environment. Thus, we discuss how the biparental QTL mapping approach has been used to dissect the genetic architecture of PUE traits. Unprecedented advances in legume genomics resources, especially high-throughput single nucleotide polymorphisms, have facilitated uncovering genomic regions related to PUE across the whole genome using a genome-wide association mapping approach. Likewise, the availability of complete genome sequence information, pangenome sequences, and the whole-genome resequencing approach have provided novel insights into structural variation, including presence/absence and copy number variations, underpinning PUE. Simultaneously, progress in functional genomics, including transcriptomics and computational biology, has facilitated the discovery of various underlying transcription factors and the corresponding downstream P signal perception genes and candidate gene(s) controlling PUE and conferring low-P tolerance in various legumes with putative function. We also discuss updated metabolomics and proteomics approaches that have improved our understanding of various gene networks, P-starvation signaling pathways, and P acquisition and utilization of molecular mechanisms important for PUE. Finally, we summarize how novel breeding schemes, including genomic selection, speed breeding, and emerging CRISPR/Cas9-based genome editing tools, will assist in designing P-use-efficient cultivars and adapting grain legumes to low-P environments. Thus, enhancing PUE by integrating various ‘omics’ approaches could improve P-acquisition and P-utilization efficiencies in various modern grain legume cultivars grown in intensive cropping systems to restrict P-fertilizer overuse and preserve the declining non-renewable global rock phosphate reservoir for securing an economical and sustainable future agriculture. • Phosphorus (P) is one of the most essential nutrients required for plant growth and development. • P fertilizers being non-renewable, judicious utilization of P fertilizer is urgently needed for sustaining legume yield for ensuring global food security. • Plants have evolved several molecular, physiological and biochemical mechanisms to sense, acquire and utilize soil P and adapt under P impoverished environments. • ‘Omics’ approach could greatly assist in unraveling the underlying genomic regions with their function contributing to improving phosphorus use efficiency in legume crops.

Morphological, palynological and transcriptomic-based SSR assessment of peony varieties adaptive in the Jiangnan region

The peony shrub belongs to the genus Paeonia in the family Paeoniaceae, is widely used in gardens and has great ornamental and medicinal value. Affected by the local high-temperature and high-humidity climate, there are few peony varieties adapted to growth in Jiangnan, China, and the genetic relationship between the varieties is unclear. In this study, simple repeat sequences (SSRs) were constructed based on transcriptomics, and 20 new SSR molecular markers suitable for peony varieties that adaptive growth in Jiangnan region were developed and characterized. A total of 144 alleles were detected in 38 peony germplasms, the average number of alleles per locus was 6.857, the effective number of alleles ranged from 2.000 to 6.191, and the average was 3.398. The observed heterozygosity (Ho) and expected heterozygosity (He) ranged from 0.056 to 1.000 and 0.258 to 0.855, respectively. Polymorphic information content (PIC) ranged from 0.243 to 0.842 with an average of 0.581, and the Shannon information index (I) ranged from 0.478 to 2.217 and was 1.318 on average. The markers exhibited good polymorphism and generality. The outer wall decoration of pollen grains and the UPGMA clustering method based on the genetic similarity coefficient were used to evaluate the genetic relationship among the peony varieties adapted to the Jiangnan region. The results showed that the results of the cluster analysis using two methods were generally consistent. Most of the native species in Jiangnan were clustered together, and the imported species were clustered together, but some cultivars could not be well distinguished by the differentiation morphology of the pollen outer wall, and the UPGMA clustering tree based on SSR molecular markers complemented them. In addition, allele-based population structure analysis further explained the genetic relationship of these varieties. The results of the population structure map showed that extensive gene exchanges occurred among the peony varieties in Jiangnan and the genetic background was more complex; the closer the genetic relationship was, the more similar the genetic composition of the cultivar. The results of this study will provide important auxiliary breeding tools and theoretical guidance for evaluating the genetic diversity, breeding practices, and germplasm innovation of peony varieties in the Jiangnan region.

WSVAS: A YOLOv4 -based phenotyping platform for automatically detecting the salt tolerance of wheat based on seed germination vigour.

Salt stress is one of the major environmental stress factors that affect and limit wheat production worldwide. Therefore, properly evaluating wheat genotypes during the germination stage could be one of the effective ways to improve yield. Currently, phenotypic identification platforms are widely used in the seed breeding process, which can improve the speed of detection compared with traditional methods. We developed the Wheat Seed Vigour Assessment System (WSVAS), which enables rapid and accurate detection of wheat seed germination using the lightweight convolutional neural network YOLOv4. The WSVAS system can automatically acquire, process and analyse image data of wheat varieties to evaluate the response of wheat seeds to salt stress under controlled environments. The WSVAS image acquisition system was set up to continuously acquire images of seeds of four wheat varieties under three types of salt stress. In this paper, we verified the accuracy of WSVAS by comparing manual scoring. The cumulative germination curves of wheat seeds of four genotypes under three salt stresses were also investigated. In this study, we compared three models, VGG16 + Faster R-CNN, ResNet50 + Faster R-CNN and YOLOv4. We found that YOLOv4 was the best model for wheat seed germination target detection, and the results showed that the model achieved an average detection accuracy (mAP) of 97.59%, a recall rate (Recall) of 97.35% and the detection speed was up to 6.82 FPS. This proved that the model could effectively detect the number of germinating seeds in wheat. In addition, the germination rate and germination index of the two indicators were highly correlated with germination vigour, indicating significant differences in salt tolerance amongst wheat varieties. WSVAS can quantify plant stress caused by salt stress and provides a powerful tool for salt-tolerant wheat breeding.

Comparative Characterization of Grain Protein Content and Composition by Chromatography-Based Separation Methods (SE-HPLC and RP-HPLC) of Ten Wheat Varieties Grown in Different Agro-Ecological Zones of Algeria

To characterize and compare the protein quality of ten durum wheat genotypes grown under three cropping modalities in Algeria (subhumid in Algiers, subhumid-semiarid in Constantine, and semiarid in Sétif), the protein profile of their kernels was performed by High-Performance Liquid Chromatography (SE-HPLC and RP-HPLC). The “variety” factor has a major impact, mainly on the insoluble fraction (Fi), on the gliadin/glutenin ratio, on the large and small glutenin aggregates (F1 and F2, respectively), and on ω-gliadins and high molecular weight albumins (F3). Conversely, the total protein content and the albumin-globulin fraction (F5) depend mainly on the environment. The α- β- and γ-gliadins (F4) are equally dependent on variety and environment. The subhumid-semiarid agroecological conditions of Constantine (SH-SA) favored an important accumulation of proteins (14.1%), particularly by an increased synthesis of omega gliadins and high-molecular-weight glutenin subunits (HMW-GS), compared to those of Algiers (SH) and Sétif (SA). For these latter environments, metabolic-type proteins are predominant, reflected in a higher F5 fraction (p < 0.05) (albumin and globulin), and significantly more alpha-beta and gamma gliadins. The use of chromatographic analyses to characterize wheat genotypes remains a reliable tool for breeding and variety promotion programs and can provide a better understanding of the ecophysiology of cereal crops.