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The Mediterranean sesame core collection contains agro-morphologically superior sesame accessions from geographically diverse regions in four continents. In the present investigation, the genetic diversity and population structure of this collection was analyzed with 5292 high-quality SNPs discovered by double-digest restriction site associated DNA (ddRAD) sequencing, a cost-effective and flexible next-generation sequencing method. The genetic distance between pairs of accessions varied from 0.023 to 0.524. The gene diversity was higher in accessions from Asia than from America, Africa, and Europe. The highest genetic differentiation was observed between accessions collected from America and Europe. Structure analysis showed the presence of three subpopulations among the sesame accessions, and only six accessions were placed in an admixture group. Phylogenetic tree and principal coordinate analysis clustered the accessions based on their countries of origin. However, no clear division was evident among the sesame accessions with regard to their continental locations. This result was supported by an AMOVA analysis, which revealed a genetic variation among continental groups of 5.53% of the total variation. The large number of SNPs clearly indicated that the Mediterranean sesame core collection is a highly diverse genetic resource. The collection can be exploited by breeders to select appropriate accessions that will provide high genetic gain in sesame improvement programs. The high-quality SNP data generated here should also be used in genome-wide association studies to explore qualitative trait loci and SNPs related to economically and agronomically important traits in sesame.

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The development and validation of different types of molecular markers is crucial to conducting marker-assisted sesame breeding. Insertion-deletion (InDel) markers are highly polymorphic and suitable for low-cost gel-based genotyping. From this perspective, this study aimed to discover and develop InDel markers through bioinformatic analysis of double digest restriction site-associated DNA sequencing (ddRADSeq) data from 95 accessions belonging to the Mediterranean sesame core collection. Bioinformatic analysis indicated the presence of 7477 InDel positions genome wide. Deletions accounted for 61% of the InDels and short deletions (1–2 bp) were the most abundant type (94.9%). On average, InDels of at least 2 bp in length had a frequency of 2.99 InDels/Mb. The 86 InDel sites having length ≥8 bp were detected in genome-wide analysis. These regions can be used for the development of InDel markers considering low-cost genotyping with agarose gels. In order to validate these InDels, a total of 38 InDel regions were selected and primers were successfully amplified. About 13% of these InDels were in the coding sequences (CDSs) and in the 3′- and 5′- untranslated regions (UTRs). Furthermore, the efficiencies of these 16 InDel markers were assessed on 32 sesame accessions. The polymorphic information content (PIC) of these 16 markers ranged from 0.06 to 0.62 (average: 0.33). These results demonstrated the success of InDel identification and marker development for sesame with the use of ddRADSeq data. These agarose-resolvable InDel markers are expected to be useful for sesame breeders.