Applied Biochemistry and Biotechnology

, Volume 170, Issue 5, pp 1163–1173 | Cite as

An Improved Micropropagation of Arnebia hispidissima (Lehm.) DC. and Assessment of Genetic Fidelity of Micropropagated Plants Using DNA-Based Molecular Markers

  • Mahendra Phulwaria
  • Manoj K. Rai
  • N. S. Shekhawat
Article
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Abstract

An efficient and improved in vitro propagation method has been developed for Arnebia hispidissima, a medicinally and pharmaceutically important plant species of arid and semiarid regions. Nodal segments (3–4 cm) with two to three nodes obtained from field grown plants were used as explants for shoot proliferation. Murashige and Skoog’s (MS) medium supplemented with cytokinins with or without indole-3-acetic acid (IAA) or naphthalene acetic acid was used for shoot multiplication. Out of different PGRs combinations, MS medium containing 0.5 mg l−1 6-benzylaminopurine and 0.1 mg l−1 IAA was optimal for shoot multiplication. On this medium, explants produced the highest number of shoots (47.50 ± 0.38). About 90 % of shoots rooted ex vitro on sterile soilrite under the greenhouse condition when the base (2–4 mm) of shoots was treated with 300 mg l−1 of indole-3-butyric acid for 5 min. The plantlets were hardened successfully in the greenhouse with 85–90 % survival rate. Random amplified polymorphic DNA (RAPD) and inter-simple sequence repeat (ISSR) markers were employed to assess the genetic stability of in vitro-regenerated plants of A. hispidissima. Out of 40 (25 RAPD and 15 ISSR) primers screened, 15 RAPD and 7 ISSR primers produced a total number of 111 (77 RAPD and 34 ISSR) reproducible amplicons. The amplified products were monomorphic across all the micropropagated plants and were similar to the mother plant. To the best of our knowledge, it is the first report on the assessment of the genetic fidelity in micropropagated plants of A. hispidissima.

Keywords

Ex vitro rooting Genetic stability Molecular markers Multiplication Shikonin 
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Notes

Acknowledgments

The authors (MP, MKR) wish to acknowledge the support of the University Grants Commission, New Delhi, for the award of Post Doctoral Fellowship and Dr. DS Kothari Post Doctoral Fellowship, respectively. We thank Department of Biotechnology, Government of India, New Delhi, for providing funds for the establishment of laboratory and greenhouse infrastructure used for the present research.

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Copyright information

© Springer Science+Business Media New York 2013

Authors and Affiliations

  • Mahendra Phulwaria
    • 1
  • Manoj K. Rai
    • 1
  • N. S. Shekhawat
    • 1
  1. 1.Biotechnology Unit, Department of BotanyJai Narain Vyas UniversityJodhpurIndia

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