Abstract
This study investigated the effects of different concentrations of zeatin and thidiazuron on the accumulation of phenolics and activity of phenylalanine ammonia lyase gene in the micropropagated Satureja spicigera shoots. S. spicigera clones were generated from cultured nodal segments on Murashige and Skoog including vitamins medium supported with zeatin and thidiazuron concentrations. Shoots were subcultured three times in the same medium and harvested at the end of the third month. The expression levels of phenylalanine ammonia lyase gene were conducted using semi-quantitative reverse transcription polymerase chain reaction technique. Rosmarinic acid contents were determined using reverse phase-high performance liquid chromatography with a diode array detector. Genetic stability of regenerants was performed using random amplified polymorphic DNA markers. Essential oils from shoots were identified by gas chromatography–mass spectrophotometry analysis. It has been detected that 0.1 mg/L thidiazuron is more effective on the node formation and shoot elongation. Contrarily, 4.0 mg/L thidiazuron caused a significant increase in shoot multiplication. The highest biomass accumulation was calculated from medium including 1.0 mg/L zeatin. According to random amplified polymorphic DNA analysis, all banding profiles from micropropagated plants were monomorphic. Gas chromatography–mass spectrophotometry analysis showed that rising concentrations of zeatin and thidiazuron increased the thymol and decreased the carvacrol content. It was also found that rosmarinic acid production increased significantly at 1.0 mg/L zeatin application. However, thidiazuron applications had no positive effect on rosmarinic acid accumulation. Comparing phenylalanine ammonia lyase gene expression and rosmarinic acid accumulation in in vitro shoots, it was found that they were correlated.
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