EXPRESSION PROFILING OF SELECTED MICRO RNAs COUPLED WITH MOLECULAR AND BIOCHEMICAL ANALYSES OF ASIATICOSIDES IN CENTELLA ASIATICA (L.)URB IN VITRO: Expression profiling of microRNAs in C. asiatica

JISHA S.; HEMANTHAKUMAR A.  S.; GOURI P. R.; SABU K. K.

doi:10.22159/ijpps.2019v11i5.32097

Authors

JISHA S. Division of Biotechnology and Bioinformatics, Jawaharlal Nehru Tropical Botanic Garden and Research Institute, Thiruvananthapuram 695562, Kerala, India
HEMANTHAKUMAR A. S. Division of Biotechnology and Bioinformatics, Jawaharlal Nehru Tropical Botanic Garden and Research Institute, Thiruvananthapuram 695562, Kerala, India
GOURI P. R. Division of Biotechnology and Bioinformatics, Jawaharlal Nehru Tropical Botanic Garden and Research Institute, Thiruvananthapuram 695562, Kerala, India
SABU K. K. Division of Biotechnology and Bioinformatics, Jawaharlal Nehru Tropical Botanic Garden and Research Institute, Thiruvananthapuram 695562, Kerala, India http://orcid.org/0000-0002-4312-1262

DOI:

https://doi.org/10.22159/ijpps.2019v11i5.32097

Keywords:

Asiaticoside, Centella asiatica, In vitro, microRNA, Real-time RT-PCR

Abstract

Objective: Centella asiatica (L.) Urb from Umbelliferae is a potential source of secondary metabolites having immense medicinal value. Asiaticoside is the major therapeutic compound. In the present study, Identification of a possible relationship between concentration/transcript level expression of asiaticoside and concentrations of growth hormones at different growth stages was observed. The current study includes molecular and biochemical evaluation of stress generated in C. asiatica at different time intervals in vitro.

Methods: The enhancement in auxin, cytokinin and final asiaticoside content were determined using immunoassay kits for auxin, cytokinin and HPLC analysis respectively. Transcript level expression at different growth phases was carried out using real-time RT-PCR. For isolation of stress-related miRNAs, reverse transcription of total RNA using miScript II RT Kit PCR System was carried out as per instructions. The differential expression of five selected miRNAs was done by Real-Time RT-PCR. The analysis of stress in vitro was done by quantification of Hydrogen Peroxide (H₂O₂), total phenolics and total antioxidants by H₂O₂assay kit, total antioxidant assay kit and Folin Ciocalteau reagent respectively. The final asiaticoside content was determined by HPLC.

Results: Differential expression of key genes involved in asiaticoside pathway showed significantly higher transcript expression, which is in correlation with the final asiaticoside content. The enhanced expression of miRNAs and the analysis of H₂O_2,total antioxidant capacity and total phenolics are suggestive of generation of oxidative stress under controlled conditions.

Conclusion: The present study shows a direct correlation between oxidative stress and transcript/phytochemical estimation of asiaticoside content under in vitro conditions.

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Author Biography

SABU K. K., Division of Biotechnology and Bioinformatics, Jawaharlal Nehru Tropical Botanic Garden and Research Institute, Thiruvananthapuram 695562, Kerala, India

Sabu is educated in the field of Plant Sciences and has working experience in plant genetics and genomics since 1995. His research interests include response of plants against environmental changes, natural variations in plant genetic resources and incorporation of genomics tools for big data analysis.

Sabu is educated in the field of Plant Sciences and has working experience in plant genetics and genomics since 1995. His research interests include response of plants against environmental changes, natural variations in plant genetic resources and incorporation of genomics tools for big data analysis. He has carried out molecular breeding in rice and instrumental in developing seven new rice varieties in Malaysia. Besides, miRNA and transcriptome analysis were conducted in cardamom to understand genes regulating plant metabolism, yield and stress response.

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