Genetic Diversity Analysis of Onion (Allium cepa) Varieties Using ISSR Markers

Abstract

Onion (Allium cepa L.) is among the most widely cultivated vegetable crops worldwide. Despite its considerable global demand and economic importance, information on the genetic diversity of onion germplasm remains limited. Assessment of genetic diversity is essential for the effective characterisation and utilisation of onion germplasm in breeding programmes. The present study was undertaken to investigate the extent of genetic variability among seven onion varieties, namely, Bhima Light Red, Bhima Shubhra, Bhima Shakti, Bhima Safed, Bhima Raj, Bhima Kiran and Bhima Red. Of the ten ISSR primers initially screened, eight produced reproducible amplification profiles. Amplified fragments were resolved by agarose gel electrophoresis and scored as present or absent, and the polymorphism information content (PIC) was calculated to assess primer informativeness. The eight primers collectively generated 40 scorable bands, of which 31 were polymorphic and 9 were monomorphic, corresponding to an overall polymorphism of 76.25%. Primers UBC-825 and UBC-812 each exhibited 100% polymorphism, with PIC values of 0.408 and 0.469, respectively, followed by UBC-823 and ISSR-01 (80% each), UBC-801 and UBC-809 (75% each), UBC-811 (60%) and UBC-880 (40%). Cluster analysis revealed that Bhima Red and Bhima Shubhra shared the highest genetic similarity, whereas Bhima Shakti and Bhima Raj formed distinct outgroups, exhibiting the greatest genetic distance from the remaining varieties. Aims: To assess the extent of genetic diversity and genetic relationships among seven onion (Allium cepa) varieties using ISSR (Inter-Simple Sequence Repeat) molecular markers Study Design: Molecular marker-based genetic diversity analysis using ISSR-PCR, with amplified products scored as binary (present/absent) data, followed by polymorphism, PIC (Polymorphism Information Content), Jaccard similarity, and UPGMA cluster analysis. Place and Duration of Study: K. K. Wagh College of Agricultural Biotechnology, Nashik, during the year 2025-2026. Methodology: Genomic DNA was isolated from seven onion varieties: Bhima Light Red, Bhima Shubhra, Bhima Shakti, Bhima Safed, Bhima Raj, Bhima Kiran, and Bhima Red using the CTAB method described by Doyle and Doyle (1987). Ten ISSR primers were initially screened, of which eight produced reproducible amplification. The amplified fragments were resolved by agarose gel electrophoresis and scored as present (1) or absent (0). Polymorphic information content (PIC) was calculated to evaluate primer informativeness. Genetic similarity was estimated using cluster analysis, which was performed using the unweighted pair-group method with arithmetic mean. Results: The eight primers generated 40 scorable bands, including 31 polymorphic and 9 monomorphic bands, yielding an overall polymorphism of 76.25%. Primers UBC-825 and UBC-812 exhibited 100% polymorphism with PIC values of 0.408 and 0.469, respectively. Pairwise Jaccard similarity coefficients ranged from 0.3235 (Bhima Light Red vs. Bhima Safed) to 0.7714 (Bhima Red vs. Bhima Safed). UPGMA cluster analysis resolved the cultivars into distinct groups, placing Bhima Red and Bhima Shubhra in proximity, while Bhima Shakti and Bhima Raj formed distinct, genetically divergent outgroups. Conclusion: Inter-Simple Sequence Repeat (ISSR) markers successfully differentiated the evaluated onion varieties and established preliminary molecular phylogenies. These findings support baseline germplasm characterisation and parental selection, though broader validation incorporating diverse genotypes and complementary marker systems remains necessary.