Assessment of genetic variability among Stevia rebaudiana Bertoni accessions cultivated in Malaysia using microsatellites

Stevia rebaudiana Bertoni (hereinafter “stevia”) is a herbaceous perennial plant of the Asteraceae family. Originating from Paraguay, stevia is considered a temperate, self-incompatible, insect-pollinated species. Recent sugar shortages in Malaysia have increased interest in stevia as an alternative. The stevia accessions available in Malaysia today are limited in number and perform poorly, emphasizing the necessity of varietal improvement. Assessment of genetic variability, diversity and intraspecific relationships is an essential step for such breeding efforts. This study aimed to reveal the genetic diversity among 21 stevia accessions, namely SRBA-1 to SRBA-21, collected from different origins utilizing 52 microsatellite DNA markers, and to identify potential markers for further marker-trait association mapping investigations. Results showed that 42 pairs of microsatellite primers (82.6%) produced polymorphic and reproducible bands. Average number of polymorphic bands ranged from 2 to 11, with an average of 5.1 alleles per locus. The total genetic diversity detected among the accessions ranged from 0.14 to 0.81, with a mean value of 0.58. Expected heterozygosity (He) and observed heterozygosity (Ho) (with mean values of 0.52 and 0.86, respectively) showed that almost all markers had deviation (Ho>He) from Hardy–Weinberg equilibrium (HWE). This result confirmed the highly outcrossing nature of this plant. The accessions evaluated revealed a wide range of variations in dry leaf weight, stevioside, rebaudioside A, rebaudioside C and total steviol glycosides. Eleven microsatellite markers with significant associations with the phenotypic traits were identified. Biosynthesis of steviol glycosides were found to be associated with the microsatellite markers SUGMS-23, SUGMS-26, SUGMS-28 and SUGMS-35. These findings can be utilised for developing marker-assisted selection procedures in stevia toward production of superior varieties for tropical environments. © 2025, University of Malaya. All rights reserved.