Steviol glycoside accumulation and expression profiling of biosynthetic pathway genes in elicited in vitro cultures of Stevia rebaudiana

Abstract

In the present study, different elicitors, viz., polyethylene glycol (PEG), alginate (ALG), chitosan (CHI), salicylic acid (SA), and yeast extract (YE), were used in in vitro shoot cultures of Stevia rebaudiana for 4 wk to decipher their effect on growth, biomass yield, and accumulation of steviol glycosides (SG), especially rebaudiosides. The highest leaf number (16.33), root number (4.67), and shoot length (3.80 cm) were observed in media supplemented with 0.5 mg/L YE, whereas maximum shoot number (2.44) and root length (1.89 cm) were recorded in 1.5 mg/L CHI and 1.0 mg/L PEG respectively. Elicitation by 1.0 mg/L YE resulted in enhanced biomass production with a maximum fresh weight of leaves (114.01 mg/plantlet), stem (90.27 mg/plantlet), and shoots (204.28 mg/plantlet). SG content was quantified through ultra-high-performance liquid chromatography (UHPLC). Stevioside (ST) content was increased by 5-fold (0.77 mg/g leaf DW) in 2.0 mg/L ALG and rebaudioside A (RA) by 7-fold (1.9 mg/g leaf DW) in 0.5 mg/L ALG. Enhancement of SG is mainly regulated by a key gene, UGT85C2, which was further validated through qRT-PCR and in silico promoter analysis. Also, cis-regulatory elements governing transcriptional regulation linked with the biosynthesis of SG were identified in key genes. The study unveils ALG as a promising elicitor to produce quality biomass with enhanced metabolite profile in S. rebaudiana.