Exploring genetic variation in saffron (Crocus sativus L.) accessions through two-locus DNA barcoding

Abstract

Saffron (Crocus sativus L.) remains among the most costly spices worldwide due to its invaluable properties. Traditional breeding methods are restricted by saffron's male sterility and clonal multiplication, which may result in genetic deterioration and make genetic diversity within saffron populations cryptic. To overcome this cryptic diversity, we conducted a comprehensive genetic analysis of sixteen saffron accessions from Morocco, France, Greece, and Italy using two universal DNA barcodes: ITS2 and trnH-psbA. Our findings unveiled significant genetic diversity among examined accessions, characterized by considerable sequence variation. This led to the identification of a notable number of haplotypes (14 and 15) with a high haplotype diversity (HD) = 0.9, signifying extensive genetic variability. Moreover, Phylogenetic and network analyses illustrated distinct genetic clusters and evolutionary relationships among saffron populations, separating our accessions from Iranian saffron. Besides, Pairwise distance and principal component analyses confirmed the unique genetic profiles of the Assif Zimer population from Morocco. Notably, the study explores environmental factors as potential contributors to genetic differences across different regions. Finally, the assessment of ITS2 secondary structures and the generation of Two-Dimensional DNA Barcodes provided unique identities to accessions and gave valuable insights into sequence variability and molecular identification. Our study contributes to a deeper understanding of saffron genetics and provides essential information for conservation projects.