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

  • Shivani Srivastava
  • Xavier A. Conlan
  • Alok Adholeya
  • David M. CahillEmail author
Original Article

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.

Keywords

Antioxidants Chemiluminescence Hairy roots Ocimum basilicum Morphotyping Rosmarinic acid 

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

Notes

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.

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflicts of interest.

Ethical approval

This article does not contain any studies with human or animal subjects.

Supplementary material

11240_2016_973_MOESM1_ESM.pdf (184 kb)
Supplementary material 1 (PDF 182 kb)
11240_2016_973_MOESM2_ESM.pdf (188 kb)
Supplementary material 2 (PDF 188 kb)
11240_2016_973_MOESM3_ESM.pdf (254 kb)
Supplementary material 3 (PDF 252 kb)
11240_2016_973_MOESM4_ESM.pdf (182 kb)
Supplementary material 4 (PDF 181 kb)

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Copyright information

© Springer Science+Business Media Dordrecht 2016

Authors and Affiliations

  • Shivani Srivastava
    • 1
    • 2
  • Xavier A. Conlan
    • 2
  • Alok Adholeya
    • 1
  • David M. Cahill
    • 2
    Email author
  1. 1.TERI–Deakin Nanobiotechnology CentreThe Energy and Resources Institute (TERI)New DelhiIndia
  2. 2.Deakin University, Geelong, Australia. Centre for Chemistry and Biotechnology, School of Life and Environmental SciencesGeelongAustralia

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