A Protocol For Synthetic Seeds From Salvia Officinalis L. Shoot Tips
Shoot tips excised from shoot culture of Salvia officinalis were encapsulated in 2% or 3% (w/v) sodium alginate and exposed to 50 mM calcium chloride for complexation. Immediately or after 6, 12 or 24 weeks of storage at 4°C, the synthetic seeds were cultured for 6 weeks on half-strength MS medium supplemented with indole-3-acetic acid (IAA) (0.1 mg/l) and solidified with 0.7% agar. The frequency of shoot and root emergence from encapsulated shoot tips was affected by the concentrations of sodium alginate and additives in the gel matrix (sucrose, gibberellic acid, MS nutrient medium) as well as duration of storage. The frequency of shoot and root induction of non-stored synthetic seeds was highest with shoot tips encapsulated with 2% sodium alginate containing 1.5% sucrose and 0.5 mg/l gibberellic acid (GA3). Shoot tips maintained their viability and ability to develop shoots even after 24 weeks of storage when they were encapsulated in 3% alginate with 1/3 MS medium, sucrose (1.5%) and GA3 (0.25 mg/l). Root formation tended to decrease with storage time. Overall, 90% of the plantlets derived from stored and non-stored synthetic seeds survived in the greenhouse and grew to phenotypically normal plants. This procedure can enable the use of synthetic seed technology for germplasm conservation of S. officinalis, a plant species of high medical and commercial value.