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

  • Medicinal Plants
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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.

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Acknowledgments

The authors are thankful to the Director, Dr. Sanjay Kumar, CSIR-IHBT for providing necessary facilities. Ashrita is thankful to DBT for providing JRF fellowship and AcSIR for Ph. D. enrolment. The authors are also thankful to Dr. Ashok Kumar and Dr. Sanatsujat Singh for providing plant materials. CSIR-IHBT publication number for this manuscript is 4486.

Funding

This study received financial support from the Council of Scientific and Industrial Research (CSIR), Government of India, under the project “Biotechnological interventions for sustainable bio-economy generation through characterization, conservation, prospection, and utilization of Himalayan bioresources (MLP-0201) and Phytopharma mission project (HCP-0010).”

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Author notes

  1. Kanika Thakur and Ashrita contributed equally and are considered as the first authors.

Authors and Affiliations

  1. Cell and Tissue Engineering Laboratory, Biotechnology Division, CSIR-Institute of Himalayan Bioresource Technology, Palampur, Himachal Pradesh, 176061, India

    Kanika Thakur,  Ashrita, Archit Sood & Ashish R. Warghat

  2. Academy of Scientific and Innovative Research(AcSIR), Ghaziabad, 201002, India

    Kanika Thakur,  Ashrita, Dinesh Kumar & Ashish R. Warghat

  3. Natural Product Chemistry and Process Development Division, CSIR-Institute of Himalayan Bioresource Technology, Palampur, Himachal Pradesh, 176061, India

    Pawan Kumar & Dinesh Kumar

Authors
  1. Kanika Thakur
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  2. Ashrita
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  3. Archit Sood
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  4. Pawan Kumar
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  6. Ashish R. Warghat
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Contributions

Conceived the concept—KT, Ashrita, and ARW. Framed the experimental design—KT, Ashrita, and ARW. Tissue culture—KT, Ashrita, and ARW. Phytochemical analysis—KT, Ashrita, PK, DK, and ARW. Gene expression profiling—Ashrita, AS, and ARW. In silico analysis: Ashrita, AS, and ARW. Data recording and statistical analysis—KT, Ashrita, and ARW. Manuscript writing (original draft)—KT, Ashrita, AS, and ARW. Manuscript writing (review and editing)—KT, Ashrita, AS, and ARW.

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Correspondence to Ashish R. Warghat.

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The authors declare that there is no conflict of interest.

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Research article followed the ethical standard of the institute.

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Editor: Yong Eui Choi

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Thakur, K., Ashrita, Sood, A. et al. Steviol glycoside accumulation and expression profiling of biosynthetic pathway genes in elicited in vitro cultures of Stevia rebaudiana. In Vitro Cell.Dev.Biol.-Plant 57, 214–224 (2021). https://doi.org/10.1007/s11627-020-10151-3

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  • DOI: https://doi.org/10.1007/s11627-020-10151-3

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