Genotyping by RAD sequencing analysis evaluated the genetic distinctiveness and relationships of chicory populations (Cichorium intybus L. var foliosum 2n=2x=18).

Cichorium intybus L. is an important horticultural leafy vegetable cultivated worldwide, as in the countries of the Mediterranean basin as in the Americas, Oceania, Africa and Southern Asia, and its phenotypic variability is well represented by the large number of local biotypes, interspecific hybrids and cultivars. For the registration and protection of new crops' commercial lines, the characterization of genetic diversity and homozygosity are fundamental. Regarding this, the Distinctness, Uniformity and Stability (DUS test) are the three majors aspects considered in the constitution, registration and protection of a new variety. To verify these parameters, several biotechnological tools and approaches can be adopted, as well as these can be for analyzing crops’ breeding populations for the selection of the best parental lines to use in planning crossings. Experimental population structure analysis and information about the genetic distinctiveness of commercial materials are essential not only for crop breeding programs, but also for protecting the commercial varieties from theft or to trace them along the food products chain to guarantee better and more appealing products to the consumers. Considering these several aims in the development and protection of crop cultivars, the purpose of our research was to assess the genetic relationships of 94 chicory samples (Chicorium intybus var foliosum), belonging to four different populations of chicory, using a genotyping-by-sequencing approach based on the Restriction site-associated DNA sequencing (RAD-seq) technology. The selected method aimed at obtaining a considerable amount of molecular data as bi-allelic single nucleotide polymorphisms (2953 SNPs) marker. The molecular information were then used to determine homozygosity (82.9% on average) and mean genetic similarity (74.0% to 87.9%), to investigate the genetic relationships among the populations considered, and to reconstruct the genetic structure of the core collection. Moreover, a further investigation was performed to identify the SNP contained in genic coding sequences (over 40%), which could be of future interest for the development of screening or selection essays. The obtained sequencing results were suitable for their bioinformatical analyses due to the absence of missing data. Combined with this, the statistical computations highlighted the genetic similarity and the genetic structure of the core collection and confirmed the distinctiveness of the analyzed populations. In this study, the RAD-Seq approach demonstrated its informativeness and reliability in the molecular characterization of breeding populations of chicory, allowing to cluster individuals according to the four populations they represented and to assess their distinctiveness and uniformity. Future studies could take place starting from this study, that could lead to the development of a standardized analytical method that gives reliable information for chicory variety breeding, registration and protection.