Elite hairy roots of Ocimum basilicum as a new source of rosmarinic acid and antioxidants
- 494 Downloads
This study reports Agrobacterium rhizogenes-mediated transformation of three cultivars of Ocimum basilicum for hairy root establishment, screening and selection for the production of rosmarinic acid and antioxidants. Hairy root development was found to be explant-specific and virulence-dependent. Distinct inter-cultivar morphological variability was found between the seven axenically developed hairy root lines and morphological traits were found to be correlated with the presence of aux2 genes, their expression and endogenous IAA content. Further inter-cultivar variability in the content of total phenolics, rosmarinic acid and caffeic acid was also found. Production of rosmarinic acid was found to be age-dependent and cultivar-specific. Chemiluminescence analysis showed the hairy roots to be rich in antioxidants and that rosmarinic acid was the major antioxidant molecule. The concentration of rosmarinic acid was found to be positively correlated with the total antioxidant potential of the hairy root extracts. On the basis of origin, morphology and metabolite content, three elite hairy root lines were selected that had significantly higher rosmarinic acid production, biomass and antioxidant potential than non-transformed roots. These new lines are rich reserves of both antioxidants and rosmarinic acid.
KeywordsAntioxidants Chemiluminescence Hairy roots Ocimum basilicum Morphotyping Rosmarinic acid
2,2-Azinobis-(3-ethylbenzothiazoline-6-sulphonic acid) radical cation
Gallic acid equivalents
High performance liquid chromatography
Indole acetic acid
Murashige and Skoog media
Modified white's medium
Naphthalene acetic acid
Ortho phosphoric acid
Polymerase chain reaction
Rotation per minute
Left subfragment of the transferred DNA
Right subfragment of the transferred DNA
Yeast Mannitol Agar
Yeast Mannitol Broth
We acknowledge Dr. Pushplata Singh for assistance with primer design and Ms. Deep Rajni for HPLC analysis. Infrastructure support provided by TERI, India and Deakin University, Australia is also duly acknowledged. Deakin University provided a postgraduate scholarship to SS.
This study was funded by Deakin University, Australia.
DC and AA conceived the work and provided comments on all drafts of the manuscript. XC provided technical expertise on total antioxidant and individual antioxidant chemiluminescence analysis. SS designed and carried out all the experiments, analyzed the results, prepared all the figures and tables and drafted the manuscript.
Compliance with ethical standards
Conflict of interest
The authors declare that they have no conflicts of interest.
This article does not contain any studies with human or animal subjects.
- Batra J, Dutta A, Singh D, Kumar S, Sen J (2004) Growth and terpenoid indole alkaloid production in Catharanthus roseus hairy root clones in relation to left- and right-termini-linked Ri T-DNA gene integration. Plant Cell Rep 23(3):148–154. doi: 10.1007/s00299-004-0815-x CrossRefPubMedGoogle Scholar
- McDermott GP, Conlan XA, Noonan LK, Costin JW, Mnatsakanyan M, Shalliker RA, Barnett NW, Francis PS (2011) Screening for antioxidants in complex matrices using high performance liquid chromatography with acidic potassium permanganate chemiluminescence detection. Anal Chim Acta 684(1–2):134–141. doi: 10.1016/j.aca.2010.10.046 CrossRefPubMedGoogle Scholar
- Nopo-Olazabal C, Hubstenberger J, Nopo-Olazabal L, Medina-Bolivar F (2013) Antioxidant activity of selected stilbenoids and their bioproduction in hairy root cultures of muscadine grape (Vitis rotundifolia Michx.). J Agric Food Chem 61(48):11744–11758. doi: 10.1021/jf400760k CrossRefPubMedGoogle Scholar
- Nopo-Olazabal C, Condori J, Nopo-Olazabal L, Medina-Bolivar F (2014) Differential induction of antioxidant stilbenoids in hairy roots of Vitis rotundifolia treated with methyl jasmonate and hydrogen peroxide. Plant Physiol Biochem 74:50–69. doi: 10.1016/j.plaphy.2013.10.035 CrossRefPubMedGoogle Scholar
- Singleton VL, Rossi JA (1965) Colorimetry of total phenolics with phosphomolybdic–phosphotungstic acid reagents. Am J Enol Vitic 16(3):144–158Google Scholar
- Thiruvengadam M, Praveen N, Maria John KM, Yang Y-S, Kim S-H, Chung I-M (2014) Establishment of Momordica charantia hairy root cultures for the production of phenolic compounds and determination of their biological activities. Plant Cell Tissue Org 118(3):545–557. doi: 10.1007/s11240-014-0506-4 CrossRefGoogle Scholar
- Xiao Y, Zhang L, Gao S, Saechao S, Di P, Chen J, Chen W (2011) The c4h, tat, hppr and hppd genes prompted engineering of rosmarinic acid biosynthetic pathway in Salvia miltiorrhiza hairy root cultures. PLoS One 6(12):e29713. doi: 10.1371/journal.pone.0029713 CrossRefPubMedPubMedCentralGoogle Scholar