CRISPR-Cas9-based repeat depletion for high-throughput genotyping of complex plant genomes.

High-throughput genotyping enables the large-scale analysis of genetic diversity in population genomics and genome-wide association studies that combine the genotypic and phenotypic characterization of large collections of accessions. Sequencing-based approaches for genotyping are progressively replacing traditional genotyping methods because of the lower ascertainment bias. However, genome-wide genotyping based on sequencing becomes expensive in species with large genomes and a high proportion of repetitive DNA. Here we describe the use of CRISPR-Cas9 technology to deplete repetitive elements in the 3.76-Gb genome of lentil (Lens culinaris), 84% consisting of repeats, thus concentrating the sequencing data on coding and regulatory regions (single-copy regions). We designed a custom set of 566,766 gRNAs targeting 2.9 Gbp of repeats and excluding repetitive regions overlapping annotated genes and putative regulatory elements based on ATAC-seq data. The novel depletion method removed ∼40% of reads mapping to repeats, increasing those mapping to single-copy regions by ∼2.6-fold. When analyzing 25 million fragments, this repeat-to-single-copy shift in the sequencing data increased the number of genotyped bases of ∼10-fold compared to nondepleted libraries. In the same condition, we were also able to identify ∼12-fold more genetic variants in the single-copy regions and increased the genotyping accuracy by rescuing thousands of heterozygous variants that otherwise would be missed because of low coverage. The method performed similarly regardless of the multiplexing level, type of library or genotypes, including different cultivars and a closely related species (L. orientalis). Our results showed that CRISPR-Cas9-driven repeat depletion focuses sequencing data on single-copy regions, thus improving high-density and genome-wide genotyping in large and repetitive genomes.

Selection and adaptive introgression guided the complex evolutionary history of the European common bean.

Domesticated crops have been disseminated by humans over vast geographic areas. Common bean (Phaseolus vulgaris L.) was introduced in Europe after 1492. Here, by combining whole-genome profiling, metabolic fingerprinting and phenotypic characterisation, we show that the first common bean cultigens successfully introduced into Europe were of Andean origin, after Francisco Pizarro's expedition to northern Peru in 1529. We reveal that hybridisation, selection and recombination have shaped the genomic diversity of the European common bean in parallel with political constraints. There is clear evidence of adaptive introgression into the Mesoamerican-derived European genotypes, with 44 Andean introgressed genomic segments shared by more than 90% of European accessions and distributed across all chromosomes except PvChr11. Genomic scans for signatures of selection highlight the role of genes relevant to flowering and environmental adaptation, suggesting that introgression has been crucial for the dissemination of this tropical crop to the temperate regions of Europe.

Towards the Development, Maintenance, and Standardized Phenotypic Characterization of Single-Seed-Descent Genetic Resources for Common Bean.

The optimal use of legume genetic resources represents a key prerequisite for coping with current agriculture-related societal challenges, including conservation of agrobiodiversity, agricultural sustainability, food security, and human health. Among legumes, the common bean (Phaseolus vulgaris) is the most economically important for human consumption, and its evolutionary trajectories as a species have been crucial to determining the structure and level of its present and available genetic diversity. Genomic advances are considerably enhancing the characterization and assessment of important genetic variants. For this purpose, the development and availability of, and access to, well-described and efficiently managed genetic resource collections that comprise pure lines derived by single-seed-descent cycles will be paramount for the use of the reservoir of common bean variability and for the advanced breeding of legume crops. This is one of the main aims of the new and challenging European project INCREASE, which is the implementation of Intelligent Collections with appropriate standardized protocols that must be characterized, maintained, and made available, along with the related data, to users such as breeders and researchers. © 2021 The Authors. Current Protocols published by Wiley Periodicals LLC. Basic Protocol 1: Characterizing common bean seeds for seed trait descriptors Basic Protocol 2: Bean seed imaging Basic Protocol 3: Characterizing bean lines for plant trait descriptors specific for common bean Primary Seed Increase.

Current State and Perspectives in Population Genomics of the Common Bean.

* Correspondence: r [...].

Adaptation to novel environments during crop diversification.

In the context of the global challenge of climate change, mitigation strategies are needed to adapt crops to novel environments. The main goal to address this is an understanding of the genetic basis of crop adaptation to different agro-ecological conditions. The movement of crops during the Colombian Exchange that started with the travels of Columbus in 1492 is an example of rapid adaptation to novel environments. Many diversification-related traits have been characterised in multiple crop species, and association-mapping analyses have identified loci involved in these. Here, we present an overview of current knowledge regarding the molecular basis related to the complex patterns of crop adaptation and dissemination, particularly outside their centres of origin. Investigation of the genomic basis of crop expansion offers a powerful contribution to the development of tools to identify and exploit valuable genetic diversity and to improve and design novel resilient crop varieties.

Phytic acid transport in Phaseolus vulgaris: A new low phytic acid mutant in the PvMRP1 gene and study of the PvMRPs promoters in two different plant systems.

Phytic acid (InsP6) is the main storage form of phosphate in seeds. In the plant it plays an important role in response to environmental stress and hormonal changes. InsP6 is a strong chelator of cations, reducing the bioavailability of essential minerals in the diet. Only a common bean low phytic acid (lpa1) mutant, affected in the PvMRP1 gene, coding for a putative tonoplastic phytic acid transporter, was described so far. This mutant is devoid of negative pleiotropic effects normally characterising lpa mutants. With the aim of isolating new common bean lpa mutants, an ethyl methane sulfonate mutagenized population was screened, resulting in the identification of an additional lpa1 allele. Other putative lpa lines were also isolated. The PvMRP2 gene is probably able to complement the phenotype of mutants affected in the PvMRP1 gene in tissues other than the seed. Only the PvMRP1 gene is expressed at appreciable levels in cotyledons. Arabidopsis thaliana and Medicago truncatula transgenic plants harbouring 1.5 kb portions of the intergenic 5' sequences of both PvMRP genes, fused upstream of the GUS reporter, were generated. GUS activity in different organs suggests a refined, species-specific mechanisms of regulation of gene expression for these two PvMRP genes.