Garlic (Allium sativum) is used worldwide in cooking and industry, including pharmacology/medicine and cosmetics, for its interesting properties. Identifying redundancies in germplasm blanks to generate core collections is a major concern, mostly in large stocks, in order to reduce space and maintenance costs. Yet, similar appearance and phenotypic plasticity of garlic varieties hinder their morphological classification. Molecular studies are challenging, due to the large and expected complex genome of this species, with asexual reproduction. Classical molecular markers, like isozymes, RAPD, SSR or AFLP, are not convenient to generate germplasm core-collections for this species. The recent emergence of high-throughput Genotyping-By-Sequencing (GBS) approaches, like DArTseq, allow to overcome such limitations to characterize and protect genetic diversity. Therefore, such technology was used in this work to: i) assess genetic diversity and structure of a large garlic-germplasm bank (417 accessions); ii) create a core collection; iii) relate genotype to agronomical features; and iv) describe a cost-effective method to manage genetic diversity in garlic-germplasm banks. Hierarchical-cluster analysis, principal-coordinates analysis and STRUCTURE showed general consistency, generating three main garlic-groups, mostly determined by variety and geographical origin. In addition, high-resolution genotyping identified 286 unique and 131 redundant accessions, used to select a reduced size germplasm-bank core collection. This demonstrates that DArTseq is a cost-effective method to analyze species with large and expected complex genomes, like garlic. To the best of our knowledge, this is the first report of high-throughput genotyping of a large garlic germplasm. This is particularly interesting for garlic adaptation and improvement, to fight biotic and abiotic stresses, in the current context of climate change and global warming.

Wheat can adapt to most agricultural conditions across temperate regions. This success is the result of phenotypic plasticity conferred by a large and complex genome composed of three homoeologous genomes (A, B, and D). Although drought is a major cause of yield and quality loss in wheat, the adaptive mechanisms and gene networks underlying drought responses in the field remain largely unknown. Here, we addressed this by utilizing an interdisciplinary approach involving field water status phenotyping, sampling, and gene expression analyses. Overall, changes at the transcriptional level were reflected in plant spectral traits amenable to field-level physiological measurements, although changes in photosynthesis-related pathways were found likely to be under more complex post-transcriptional control. Examining homoeologous genes with a 1:1:1 relationship across the A, B, and D genomes (triads), we revealed a complex genomic architecture for drought responses under field conditions, involving gene homoeolog specialization, multiple gene clusters, gene families, miRNAs, and transcription factors coordinating these responses. Our results provide a new focus for genomics-assisted breeding of drought-tolerant wheat cultivars.

Climatic conditions affect the growth, development and final crop production. As wheat is of paramount importance as a staple crop in the human diet, there is a growing need to study its abiotic stress adaptation through the performance of key breeding traits. New and complementary approaches, such as genome-wide association studies (GWAS) and genomic selection (GS), are used for the dissection of different agronomic traits. The present study focused on the dissection of agronomic and quality traits of interest (initial agronomic score, yield, gluten index, sedimentation index, specific weight, whole grain protein and yellow colour) assessed in a panel of 179 durum wheat lines (Triticum durum Desf.), grown under rainfed conditions in different Mediterranean environments in Southern Spain (Andalusia). The findings show a total of 37 marker-trait associations (MTAs) which affect phenotype expression for three quality traits (specific weight, gluten and sedimentation indexes). MTAs could be mapped on the A and B durum wheat subgenomes (on chromosomes 1A, 1B, 2A, 2B and 3A) through the recently available bread wheat reference assembly (IWGSC RefSeqv1). Two of the MTAs found for quality traits (gluten index and SDS) corresponded to the known Glu-B1 and Glu-A1 loci, for which candidate genes corresponding to high molecular weight glutenin subunits could be located. The GS prediction ability values obtained from the breeding materials analyzed showed promising results for traits as grain protein content, sedimentation and gluten indexes, which can be used in plant breeding programs.

Final grain production and quality in durum wheat are affected by biotic and abiotic stresses. The association mapping (AM) approach is useful for dissecting the genetic control of quantitative traits, with the aim of increasing final wheat production under stress conditions. In this study, we used AM analyses to detect quantitative trait loci (QTL) underlying agronomic and quality traits in a collection of 294 elite durum wheat lines from CIMMYT (International Maize and Wheat Improvement Center), grown under different water regimes over four growing seasons. Thirty-seven significant marker-trait associations (MTAs) were detected for sedimentation volume (SV) and thousand kernel weight (TKW), located on chromosomes 1B and 2A, respectively. The QTL loci found were then confirmed with several AM analyses, which revealed 12 sedimentation index (SDS) MTAs and two additional loci for SV (4A) and yellow rust (1B). A candidate gene analysis of the identified genomic regions detected a cluster of 25 genes encoding blue copper proteins in chromosome 1B, with homoeologs in the two durum wheat subgenomes, and an ubiquinone biosynthesis O-methyltransferase gene. On chromosome 2A, several genes related to photosynthetic processes and metabolic pathways were found in proximity to the markers associated with TKW. These results are of potential use for subsequent application in marker-assisted durum wheat-breeding programs.

The practical use of molecular markers is facilitated by cost-effective detection techniques. In this work, wheat insertion site-based polymorphisms (ISBP) markers were set up for genotyping using high-resolution melting analysis (HRM). Polymorphic HRM-ISBP assays were developed for wheat chromosomes 4A and 3B and used for wheat variability assessment. The marker sequences were mapped against the wheat genome reference sequence, targeting interesting genes. Those genes were located within or in proximity to previously described quantitative trait loci (QTL) or meta-quantitative trait loci (MQTL) for drought and heat stress tolerance, and also yield and yield related traits. Eighteen of the markers used tagged drought related genes and, interestingly, eight of the genes were differentially expressed under different abiotic stress conditions. These results confirmed HRM as a cost-effective and efficient tool for wheat breeding programs.

High-resolution melting analysis (HRM) is a resolutive technique, using PCR amplification and in-tube detection, which is based on the PCR product’s melting analysis. It is a promising technique for breeding analysis, as it does not require dedicated sequencing equipment. It can be performed using QRT-PCR equipment that can be available in small-medium molecular biology laboratories or locally by the breeders, and it does not require an electrophoretic step to analyze the amplified DNA fragments. To develop effective HRM assays, the search for highly polymorphic sites amenable to PCR amplification is a prerequisite, which is not an easy task in wheat due to its genome complexity. The insertion site-based polymorphism markers (ISBPs) are PCR markers designed based on the knowledge of the sequence flanking transposable element (TE) sequences. The two PCR primers are designed with one in the transposable element and the other in the flanking DNA sequence. TEs are very abundant and nested in the wheat genome, with unique (genome-specific) insertion sites that are highly polymorphic. In this work, we analyze the available HRM-ISBP assays for wheat 3B and 4A chromosomes, and update their applications in wheat diversity at drought and heat MQTL loci.