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Elite hairy roots of Ocimum basilicum as a new source of rosmarinic acid and antioxidants

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Abstract

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.

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Abbreviations

ABTS·+ :

2,2-Azinobis-(3-ethylbenzothiazoline-6-sulphonic acid) radical cation

CA:

Caffeic acid

dNTPs:

Nucleoside triphosphate

DPPH· :

2,2-Diphenyl-1-picrylhydrazyl radical

GAE:

Gallic acid equivalents

HPLC:

High performance liquid chromatography

HR:

Hairy root

IAA:

Indole acetic acid

M:

Minimal medium

MS:

Murashige and Skoog media

MW:

Modified white's medium

NAA:

Naphthalene acetic acid

NAM:

Naphthalene acetamide

OPA:

Ortho phosphoric acid

PCR:

Polymerase chain reaction

RA:

Rosmarinic acid

rpm:

Rotation per minute

Smr:

Streptomycin resistant

T-DNA:

Transferred DNA

TL-DNA:

Left subfragment of the transferred DNA

TR-DNA:

Right subfragment of the transferred DNA

YMA:

Yeast Mannitol Agar

YMB:

Yeast Mannitol Broth

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Acknowledgments

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.

Funding

This study was funded by Deakin University, Australia.

Author’s contribution

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.

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Authors and Affiliations

  1. TERI–Deakin Nanobiotechnology Centre, The Energy and Resources Institute (TERI), DS Block, India Habitat Centre, Lodhi Road, New Delhi, 110003, India

    Shivani Srivastava & Alok Adholeya

  2. Deakin University, Geelong, Australia. Centre for Chemistry and Biotechnology, School of Life and Environmental Sciences, (Waurn Ponds Campus), 75 Pigdons Road, Locked Bag 20000, Geelong, VIC, 3220, Australia

    Shivani Srivastava, Xavier A. Conlan & David M. Cahill

Authors
  1. Shivani Srivastava
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  2. Xavier A. Conlan
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  3. Alok Adholeya
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  4. David M. Cahill
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Correspondence to David M. Cahill.

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Srivastava, S., Conlan, X.A., Adholeya, A. et al. Elite hairy roots of Ocimum basilicum as a new source of rosmarinic acid and antioxidants. Plant Cell Tiss Organ Cult 126, 19–32 (2016). https://doi.org/10.1007/s11240-016-0973-x

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  • DOI: https://doi.org/10.1007/s11240-016-0973-x

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