Planta Med 2016; 82 - PB23
DOI: 10.1055/s-0036-1578671

Genetic Fidelity Of Stevia Rebaudiana Bertoni Plants Grown From Synthetic Seeds Following In Vitro Storage

H Lata 1, S Chandra 1, N Techen 1, YH Wang 1, MA ElSohly 1, 2, IA Khan 1, 2
  • 1National Center for Natural Products Research, School of Pharmacy, University of Mississippi, MS 38677, USA
  • 2Department of Biomolecular Sciences, School of Pharmacy, University of Mississippi, MS 38677, USA

The genetic stability of Stevia rebaudiana Bertoni plants, propagated through in vitro stored synthetic seeds was assessed. In vitro grown micro shoots of Stevia rebaudiana were encapsulated in 5% sodium alginate and 50 mM CaCl2. The synthetic seeds were stored in vitro for 4 months and re-grown under the tissue culture conditions (16-h photoperiod, 25 oC) on Murashige and Skoog (MS) medium supplemented with thidiazuron (TDZ 0.2 mg L-1) [1]. Well rooted plantlets were successfully transferred to a climatic controlled indoor cultivation facility for further cultivation with mother plant. Leaf samples were taken from nine, randomly selected, fully developed, healthy in vitro propagated plants and a mother plant for the molecular analysis of genetic fidelity using eight inter simple sequence repeat (ISSR) markers. Each tested primer produced clear and scorable amplification products ranging in size from about 176 bp (UBC 817) to 1354 bp (UBC 826) with an average of 5.1 products per primer. A total of 41 bands were generated, giving rise to monomorphic patterns across all 10 plantlets analyzed. No ISSR polymorphism was observed in the micropropagated plants. Furthermore, leaf samples from the plants used for molecular analysis were also evaluated for rebaudioside A and stevioside content using high performance liquid chromatography (HPLC) [2]. Our data show no significant differences (p < 0.05) in rebaudioside A and stevioside content among re-grown plants and the mother plant following four months of in vitro storage, thus confirming the clonal fidelity of Stevia rebaudiana plants.

Acknowledgement: This research was partially funded by a grant from the USDA, Agricultural Research Service Specific Cooperative Agreement No. 58 – 6408 – 1-603 – 07.

References: [1] Hemant Lata, Suman Chandra et al. (2013) American Journal of Plant Sciences, 4: 117 – 128. [2] Yan-Hong Wang, Bharathi Avula et al. (2015) Food Additives & Contaminants: Part A, 32: 674 – 685.