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The influence of Agrobacterium rhizogenes on induction of hairy roots and ß-carboline alkaloids production in Tribulus terrestris L.

Physiology and Molecular Biology of Plants volume 20, pages 69–80 (2014)Cite this article

Abstract

We have developed an efficient transformation system for Tribulus terrestris L., an important medicinal plant, using Agrobacterium rhizogenes strains AR15834 and GMI9534 to generate hairy roots. Hairy roots were formed directly from the cut edges of leaf explants 10–14 days after inoculation with the Agrobacterium with highest frequency transformation being 49 %, which was achieved using Agrobacterium rhizogenes AR15834 on hormone-free MS medium after 28 days inoculation. PCR analysis showed that rolB genes of Ri plasmid of A. rhizogenes were integrated and expressed into the genome of transformed hairy roots. Isolated transgenic hairy roots grew rapidly on MS medium supplemented with indole-3-butyric acid. They showed characteristics of transformed roots such as fast growth and high lateral branching in comparison with untransformed roots. Isolated control and transgenic hairy roots grown in liquid medium containing IBA were analyzed to detect ß-carboline alkaloids by High Performance Thin Layer Chromatograghy (HPTLC). Harmine content was estimated to be 1.7 μg g−1 of the dried weight of transgenic hairy root cultures at the end of 50 days of culturing. The transformed roots induced by AR15834 strain, spontaneously, dedifferentiated as callus on MS medium without hormone. Optimum callus induction and shoot regeneration of transformed roots in vitro was achieved on MS medium containing 0.4 mg L−1 naphthaleneacetic acid and 2 mg L−1 6-benzylaminopurine (BAP) after 50 days. The main objective of this investigation was to establish hairy roots in this plant by using A. rhizogenes to synthesize secondary products at levels comparable to the wild-type roots.

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Abbreviations

BAP:

6-Benzylaminopurine

IBA:

Indol-3-Butyric Acid

NAA:

Naphthaleneacetic Acid

rolB :

Root locus B

PCR:

Polymerase Chain Reaction

Rf :

Retardation factor

HPTLC:

High Performance Thin Layer Chromatograghy

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Affiliations

  1. Department of Biology, Science and Research Branch, Islamic Azad University, Tehran, Iran

    Sara Sharifi & Taher Nejad Sattari

  2. Department of Plant Breeding and Agronomy, Faculty of Agriculture, Razi University, Kermanshah, Iran

    Alireza Zebarjadi

  3. Department of Biotechnology for Drought Resistance, Razi University, Kermanshah, Iran

    Alireza Zebarjadi

  4. Department of Biology, Tehran North Branch, Islamic Azad University, Tehran, Iran

    Ahmad Majd

  5. Department of Biotechnology and Chemical engineering, Science and Research Branch, Islamic Azad University, Kermanshah, Islamic Republic of Iran, Iran

    Hamidreza Ghasempour

Authors
  1. Sara Sharifi
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  2. Taher Nejad Sattari
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  3. Alireza Zebarjadi
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  4. Ahmad Majd
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  5. Hamidreza Ghasempour
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Correspondence to Sara Sharifi.

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Sharifi, S., Sattari, T.N., Zebarjadi, A. et al. The influence of Agrobacterium rhizogenes on induction of hairy roots and ß-carboline alkaloids production in Tribulus terrestris L.. Physiol Mol Biol Plants 20, 69–80 (2014). https://doi.org/10.1007/s12298-013-0208-0

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Keywords

  • Agrobacterium rhizogenes
  • Hairy roots
  • Harmine
  • Secondary metabolite
  • Transformation
  • Tribulus terrestris