Siam orange (Citrus nobilis Lour.) is a major horticultural commodity with wide distribution and the largest share in Citrus production in Indonesia. The information on genetic relationships among C. nobilis populations from major production centers remains limited in Indonesia. This study evaluated the genetic diversity, haplotype structure, and phylogenetic relationships of C. nobilis populations from four major production centers in Sumatra (Gunung Omeh, Pasaman, Bangkinang, and Berastagi) using ITS markers. Forty DNA samples were amplified using ITS1 and ITS4 primers, generating ITS (internal transcribed spacer) fragments of 603–666 bp. The resulting sequences incurred analysis for nucleotide composition, genetic diversity, haplotype structure, and phylogenetic relationships. The results revealed eight polymorphic nucleotide sites and a high GC content (65.24%). Eight haplotypes identified had the Hap_1, representing 65% of all samples and occurring across all populations. Haplotype diversity was the highest in Gunung Omeh and lowest in Berastagi. The haplotype network positioned Hap_1 as the central node, while other haplotypes differed by one to three nucleotide substitutions. This study provides the first ITS-based assessment of genetic relationships among major C. nobilis populations in Sumatra and complements previous random amplified polymorphic DNA (RAPD)-based findings with sequence-level evidence. The genetic information obtained may support germplasm selection, conservation efforts, and future breeding programs for the Siam orange in Indonesia.

Keywords: Siam orange (Citrus nobilis), genetic variation, ITS marker, haplotype diversity, phylogenetic analysis, Sumatra

Key findings: The analysis of C. nobilis from four production centers using ITS markers revealed eight haplotypes with low nucleotide diversity. The shared and region-specific haplotypes disclosed an extensive exchange of planting materials among the production centers. Phylogenetic analysis placed all accessions within a single monophyletic clade, reflecting a close genetic relationship.