Genetic diversity and population structure analysis of a diverse panel of pea (Pisum sativum).

Breeding resilient cultivars with increased tolerance to environmental stress and enhanced resistance to pests and diseases demands pre-breeding efforts that include understanding genetic diversity. This study aimed to evaluate the genetic diversity and population structure of 265 pea accessions. The diversity arrays technology (DArT) genotyping method was employed to identify single-nucleotide polymorphisms (SNPs) and silico markers. After stringent filtering, 6966 SNP and 8,454 silico markers were selected for diversity analysis. Genetic diversity was estimated by grouping accessions based on plant material type, geographic origin, growth habit, and seed color. Generally, diversity estimations obtained using SNPs were similar to those estimated using silico markers. The polymorphism information content (PIC) of the SNP markers ranged from 0.0 to 0.5, with a quarter of them displaying PIC values exceeding 0.4, making them highly informative. Analysis based on plant material type revealed narrow observed heterozygosity (Ho = 0.02-0.03) and expected heterozygosity (He = 0.26-0.31), with landrace accessions exhibiting the highest diversity. Geographic origin-based diversity analysis revealed Ho = 0.02-0.03 and He = 0.22 to 0.30, with European accessions showing the greatest diversity. Moreover, private alleles unique to landrace (4) and European (22) accessions were also identified, which merit further investigation for their potential association with desirable traits. The analysis of molecular variance revealed a highly significant genetic differentiation among accession groups classified by seed color, growth habit, plant material types, and geographic origin (p < 0.01). Principal coordinate analysis and neighbor-joining cluster analysis revealed weak clustering of accessions at different grouping levels. This study underscores the significance of genetic diversity in pea collections, offering valuable insights for targeted breeding and conservation efforts. By leveraging genomic data and exploring untapped genetic resources, pea breeding programs can be fortified to ensure sustainable plant protein production and address future challenges in agriculture.

Finger millet RNA-seq reveals differential gene expression associated with tolerance to aluminum toxicity and provides novel genomic resources.

Eleusine coracana, finger millet, is a multipurpose crop cultivated in arid and semi-arid regions of Africa and Asia. RNA sequencing (RNA-seq) was used in this study to obtain valuable genomic resources and identify genes differentially expressed between Al-tolerant and Al-susceptible genotypes. Two groups of finger millet genotypes were used: Al-tolerant (215836, 215845, and 229722) and Al-susceptible (212462, 215804 and 238323). The analysis of the RNA-seq data resulted in 198,546 unigenes, 56.5% of which were annotated with significant hits in one or more of the following six databases: NR (48.8%), GO (29.7%), KEGG (45%), PlantTFDB (19.0%), Uniprot (49.2%), and NT (46.2%). It is noteworthy that only 220 unigenes in the NR database had significant hits against finger millet sequences suggesting that finger millet's genomic resources are scarce. The gene expression analysis revealed that 322 genes were significantly differentially expressed between the Al-tolerant and Al-susceptible genotypes, of which 40.7% were upregulated while 59.3% were downregulated in Al-tolerant genotypes. Among the significant DEGs, 54.7% were annotated in the GO database with the top hits being ATP binding (GO:0005524) and DNA binding (GO:0003677) in the molecular function, DNA integration (GO:0015074) and cell redox homeostasis in the biological process, as well as cellular anatomical entity and intracellular component in the cellular component GO classes. Several of the annotated DEGs were significantly enriched for their corresponding GO terms. The KEGG pathway analysis resulted in 60 DEGs that were annotated with different pathway classes, of which carbohydrate metabolism and signal transduction were the most prominent. The homologs of a number of significant DEGs have been previously reported as being associated with Al or other abiotic stress responses in various crops, including carboxypeptidase SOL1, HMA3, AP2, bZIP, C3H, and WRKY TF genes. A more detailed investigation of these and other DEGs will enable genomic-led breeding for Al tolerance in finger millet. RNA-seq data analysis also yielded 119,073 SNP markers, the majority of which had PIC values above 0.3, indicating that they are highly informative. Additionally, 3,553 single-copy SSR markers were identified, of which trinucleotide SSRs were the most prevalent. These genomic resources contribute substantially to the enrichment of genomic databases for finger millet, and facilitate future research on this crop.

Novel GBS-Based SNP Markers for Finger Millet and Their Use in Genetic Diversity Analyses.

Eleusine coracana (L.) Gaertn., commonly known as finger millet, is a multipurpose crop used for food and feed. Genomic tools are required for the characterization of crop gene pools and their genomics-led breeding. High-throughput sequencing-based characterization of finger millet germplasm representing diverse agro-ecologies was considered an effective method for determining its genetic diversity, thereby suggesting potential candidates for breeding. In this study, the genotyping-by-sequencing (GBS) method was used to simultaneously identify novel single nucleotide polymorphism (SNP) markers and genotype 288 finger millet accessions collected from Ethiopia and Zimbabwe. The accessions were characterized at individual and group levels using 5,226 bi-allelic SNPs, with a minimum allele frequency (MAF) of above 0.05, distributed across 2,500 scaffolds of the finger millet reference genome. The polymorphism information content (PIC) of the SNPs was 0.23 on average, and a quarter of them have PIC values over 0.32, making them highly informative. The grouping of the 288 accessions into seven populations based on geographic proximity and the potential for germplasm exchange revealed a narrow range of observed heterozygosity (Ho; 0.09-0.11) and expected heterozygosity (He) that ranged over twofold, from 0.11 to 0.26. Alleles unique to the different groups were also identified, which merit further investigation for their potential association with desirable traits. The analysis of molecular variance (AMOVA) revealed a highly significant genetic differentiation among groups of accessions classified based on the geographic region, country of origin, days to flowering, panicle type, and Al tolerance (p < 0.01). The high genetic differentiation between Ethiopian and Zimbabwean accessions was evident in the AMOVA, cluster, principal coordinate, and population structure analyses. The level of genetic diversity of finger millet accessions varies moderately among locations within Ethiopia, with accessions from the northern region having the lowest level. In the neighbor-joining cluster analysis, most of the improved cultivars included in this study were closely clustered, probably because they were developed using genetically less diverse germplasm and/or selected for similar traits, such as grain yield. The recombination of alleles via crossbreeding genetically distinct accessions from different regions of the two countries can potentially lead to the development of superior cultivars.

Novel Expressed Sequence Tag-Derived and Other Genomic Simple Sequence Repeat Markers Revealed Genetic Diversity in Ethiopian Finger Millet Landrace Populations and Cultivars.

Finger millet (Eleusine coracana (L.) Geartn.) is a self-pollinating amphidiploid crop cultivated with minimal input for food and feed, as well as a source of income for small-scale farmers. To efficiently assess its genetic diversity for conservation and use in breeding programs, polymorphic DNA markers that represent its complex tetraploid genome have to be developed and used. In this study, 13 new expressed sequence tag-derived simple sequence repeat (EST-SSR) markers were developed based on publicly available finger millet ESTs. Using 10 polymorphic SSR markers (3 genomic and 7 novel EST-derived), the genetic diversity of 55 landrace accessions and 5 cultivars of finger millet representing its major growing areas in Ethiopia was assessed. In total, 26 alleles were detected across the 10 loci, and the average observed number of alleles per locus was 5.6. The polymorphic information content (PIC) of the loci ranged from 0.045 (Elco-48) to 0.71 (UGEP-66). The level of genetic diversity did not differ much between the accessions with the mean gene diversity estimates ranging only from 0.44 (accession 216054) to 0.68 (accession 237443). Similarly, a narrow range of variation was recorded at the level of regional states ranging from 0.54 (Oromia) to 0.59 (Amhara and Tigray). Interestingly, the average gene diversity of the landrace accessions (0.57) was similar to that of the cultivars (0.58). The analysis of molecular variance (AMOVA) revealed significant genetic variation both within and among accessions. The variation among the accessions accounted for 18.8% of the total variation (F ST = 0.19; P < 0.001). Similarly, significant genetic variation was obtained among the geographic regions, accounting for 6.9% of the total variation (P < 0.001). The results of the cluster, principal coordinate, and population structure analyses suggest a poor correlation between the genetic makeups of finger millet landrace populations and their geographic regions of origin, which in turn suggests strong gene flow between populations within and across geographic regions. This study contributed novel EST-SSR markers for their various applications, and those that were monomorphic should be tested in more diverse finger millet genetic resources.